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Sample records for altered host cell

  1. Inhibition of growth and alteration of host cell interactions of Pasteurella multocida with natural byproducts.

    Science.gov (United States)

    Salaheen, S; Almario, J A; Biswas, D

    2014-06-01

    Pasteurella multocida is a leading cause of fowl cholera in both free-range pasture and conventional/commercially raised poultry. Its infection is a serious threat to poultry health and overall flock viability. Organic poultry is comparatively more vulnerable to this pathogen. It is a significant cause of production loss and price increase of poultry products, specifically organic poultry products. Some plant products are well documented as sources of natural antimicrobials such as polyphenols found in different berry pomaces and citrus oil. Pomace, a byproduct (primarily of seeds and skins) of fruits used for juice and wine production, and citrus oil, the byproduct of citrus juice production, show promising antimicrobial activity against various pathogens. Here, we showed for the first time that blackberry and blueberry pomace extracts and citrus oil inhibited P. multocida growth. Minimum bactericidal concentrations were determined as 0.3 and 0.4 mg/mL gallic acid equivalent for blackberry and blueberry pomace extracts, respectively. Similarly, only 0.05% citrus oil (vol/vol) completely inhibited P. multocida growth. Under shaking conditions, the antimicrobial activity of both pomace extracts and citrus oil was more intensive. Even citrus oil vapor also significantly reduced the growth of P. multocida. In addition, cell surface hydrophobicity of P. multocida was increased by 2- to 3-fold and its adherence to chicken fibroblast (DF1) and bovine mammary gland (MacT) cells was reduced significantly in the presence of pomace extracts only. This study indicates that these natural products might be good alternatives to conventional antimicrobial agents, and hence, may be used as feed or water supplements to control fowl cholera and reduce production loss caused by P. multocida. PMID:24879687

  2. NADPH Oxidase 1 Is Associated with Altered Host Survival and T Cell Phenotypes after Influenza A Virus Infection in Mice.

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    Amelia R Hofstetter

    Full Text Available The role of the reactive oxygen species-producing NADPH oxidase family of enzymes in the pathology of influenza A virus infection remains enigmatic. Previous reports implicated NADPH oxidase 2 in influenza A virus-induced inflammation. In contrast, NADPH oxidase 1 (Nox1 was reported to decrease inflammation in mice within 7 days post-influenza A virus infection. However, the effect of NADPH oxidase 1 on lethality and adaptive immunity after influenza A virus challenge has not been explored. Here we report improved survival and decreased morbidity in mice with catalytically inactive NADPH oxidase 1 (Nox1*/Y compared with controls after challenge with A/PR/8/34 influenza A virus. While changes in lung inflammation were not obvious between Nox1*/Y and control mice, we observed alterations in the T cell response to influenza A virus by day 15 post-infection, including increased interleukin-7 receptor-expressing virus-specific CD8+ T cells in lungs and draining lymph nodes of Nox1*/Y, and increased cytokine-producing T cells in lungs and spleen. Furthermore, a greater percentage of conventional and interstitial dendritic cells from Nox1*/Y draining lymph nodes expressed the co-stimulatory ligand CD40 within 6 days post-infection. Results indicate that NADPH oxidase 1 modulates the innate and adaptive cellular immune response to influenza virus infection, while also playing a role in host survival. Results suggest that NADPH oxidase 1 inhibitors may be beneficial as adjunct therapeutics during acute influenza infection.

  3. Toxoplasma gondii is dependent on glutamine and alters migratory profile of infected host bone marrow derived immune cells through SNAT2 and CXCR4 pathways.

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    I-Ping Lee

    Full Text Available The obligate intracellular parasite, Toxoplasma gondii, disseminates through its host inside infected immune cells. We hypothesize that parasite nutrient requirements lead to manipulation of migratory properties of the immune cell. We demonstrate that 1 T. gondii relies on glutamine for optimal infection, replication and viability, and 2 T. gondii-infected bone marrow-derived dendritic cells (DCs display both "hypermotility" and "enhanced migration" to an elevated glutamine gradient in vitro. We show that glutamine uptake by the sodium-dependent neutral amino acid transporter 2 (SNAT2 is required for this enhanced migration. SNAT2 transport of glutamine is also a significant factor in the induction of migration by the small cytokine stromal cell-derived factor-1 (SDF-1 in uninfected DCs. Blocking both SNAT2 and C-X-C chemokine receptor 4 (CXCR4; the unique receptor for SDF-1 blocks hypermotility and the enhanced migration in T. gondii-infected DCs. Changes in host cell protein expression following T. gondii infection may explain the altered migratory phenotype; we observed an increase of CD80 and unchanged protein level of CXCR4 in both T. gondii-infected and lipopolysaccharide (LPS-stimulated DCs. However, unlike activated DCs, SNAT2 expression in the cytosol of infected cells was also unchanged. Thus, our results suggest an important role of glutamine transport via SNAT2 in immune cell migration and a possible interaction between SNAT2 and CXCR4, by which T. gondii manipulates host cell motility.

  4. Host genetic background influences the response to the opportunistic Pseudomonas aeruginosa infection altering cell-mediated immunity and bacterial replication.

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    Maura De Simone

    Full Text Available Pseudomonas aeruginosa is a common cause of healthcare-associated infections including pneumonia, bloodstream, urinary tract, and surgical site infections. The clinical outcome of P. aeruginosa infections may be extremely variable among individuals at risk and patients affected by cystic fibrosis. However, risk factors for P. aeruginosa infection remain largely unknown. To identify and track the host factors influencing P. aeruginosa lung infections, inbred immunocompetent mouse strains were screened in a pneumonia model system. A/J, BALB/cJ, BALB/cAnNCrl, BALB/cByJ, C3H/HeOuJ, C57BL/6J, C57BL/6NCrl, DBA/2J, and 129S2/SvPasCRL mice were infected with P. aeruginosa clinical strain and monitored for body weight and mortality up to seven days. The most deviant survival phenotypes were observed for A/J, 129S2/SvPasCRL and DBA/2J showing high susceptibility while BALB/cAnNCrl and C3H/HeOuJ showing more resistance to P. aeruginosa infection. Next, one of the most susceptible and resistant mouse strains were characterized for their deviant clinical and immunological phenotype by scoring bacterial count, cell-mediated immunity, cytokines and chemokines profile and lung pathology in an early time course. Susceptible A/J mice showed significantly higher bacterial burden, higher cytokines and chemokines levels but lower leukocyte recruitment, particularly neutrophils, when compared to C3H/HeOuJ resistant mice. Pathologic scores showed lower inflammatory severity, reduced intraluminal and interstitial inflammation extent, bronchial and parenchymal involvement and diminished alveolar damage in the lungs of A/J when compared to C3H/HeOuJ. Our findings indicate that during an early phase of infection a prompt inflammatory response in the airways set the conditions for a non-permissive environment to P. aeruginosa replication and lock the spread to other organs. Host gene(s may have a role in the reduction of cell-mediated immunity playing a critical role in

  5. Host genetic background influences the response to the opportunistic Pseudomonas aeruginosa infection altering cell-mediated immunity and bacterial replication.

    Science.gov (United States)

    De Simone, Maura; Spagnuolo, Lorenza; Lorè, Nicola Ivan; Rossi, Giacomo; Cigana, Cristina; De Fino, Ida; Iraqi, Fuad A; Bragonzi, Alessandra

    2014-01-01

    Pseudomonas aeruginosa is a common cause of healthcare-associated infections including pneumonia, bloodstream, urinary tract, and surgical site infections. The clinical outcome of P. aeruginosa infections may be extremely variable among individuals at risk and patients affected by cystic fibrosis. However, risk factors for P. aeruginosa infection remain largely unknown. To identify and track the host factors influencing P. aeruginosa lung infections, inbred immunocompetent mouse strains were screened in a pneumonia model system. A/J, BALB/cJ, BALB/cAnNCrl, BALB/cByJ, C3H/HeOuJ, C57BL/6J, C57BL/6NCrl, DBA/2J, and 129S2/SvPasCRL mice were infected with P. aeruginosa clinical strain and monitored for body weight and mortality up to seven days. The most deviant survival phenotypes were observed for A/J, 129S2/SvPasCRL and DBA/2J showing high susceptibility while BALB/cAnNCrl and C3H/HeOuJ showing more resistance to P. aeruginosa infection. Next, one of the most susceptible and resistant mouse strains were characterized for their deviant clinical and immunological phenotype by scoring bacterial count, cell-mediated immunity, cytokines and chemokines profile and lung pathology in an early time course. Susceptible A/J mice showed significantly higher bacterial burden, higher cytokines and chemokines levels but lower leukocyte recruitment, particularly neutrophils, when compared to C3H/HeOuJ resistant mice. Pathologic scores showed lower inflammatory severity, reduced intraluminal and interstitial inflammation extent, bronchial and parenchymal involvement and diminished alveolar damage in the lungs of A/J when compared to C3H/HeOuJ. Our findings indicate that during an early phase of infection a prompt inflammatory response in the airways set the conditions for a non-permissive environment to P. aeruginosa replication and lock the spread to other organs. Host gene(s) may have a role in the reduction of cell-mediated immunity playing a critical role in the control of P

  6. Adenovirus-induced alterations in host cell gene expression prior to the onset of viral gene expression.

    Science.gov (United States)

    Granberg, Fredrik; Svensson, Catharina; Pettersson, Ulf; Zhao, Hongxing

    2006-09-15

    In this report, we have studied gene expression profiles in human primary lung fibroblasts (IMR-90) during the very early phase of an adenovirus infection. Eight out of twelve genes with known functions encoded transcription factors linked to two major cellular processes; inhibition of cell growth (ATF3, ATF4, KLF4, KLF6 and ELK3) and immune response (NR4A1 and CEBPB), indicating that the earliest consequences of an adenovirus infection are growth arrest and induction of an immune response. A time course analysis showed that the induction of these immediate-early response genes was transient and suppressed after the onset of the adenovirus early gene expression. PMID:16860366

  7. Host stress hormones alter vector feeding preferences, success, and productivity.

    Science.gov (United States)

    Gervasi, Stephanie S; Burkett-Cadena, Nathan; Burgan, Sarah C; Schrey, Aaron W; Hassan, Hassan K; Unnasch, Thomas R; Martin, Lynn B

    2016-08-17

    Stress hormones might represent a key link between individual-level infection outcome, population-level parasite transmission, and zoonotic disease risk. Although the effects of stress on immunity are well known, stress hormones could also affect host-vector interactions via modification of host behaviours or vector-feeding patterns and subsequent reproductive success. Here, we experimentally manipulated songbird stress hormones and examined subsequent feeding preferences, feeding success, and productivity of mosquito vectors in addition to defensive behaviours of hosts. Despite being more defensive, birds with elevated stress hormone concentrations were approximately twice as likely to be fed on by mosquitoes compared to control birds. Moreover, stress hormones altered the relationship between the timing of laying and clutch size in blood-fed mosquitoes. Our results suggest that host stress could affect the transmission dynamics of vector-borne parasites via multiple pathways. PMID:27512147

  8. Altering host resistance to infections through microbial transplantation.

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    Benjamin P Willing

    Full Text Available Host resistance to bacterial infections is thought to be dictated by host genetic factors. Infections by the natural murine enteric pathogen Citrobacter rodentium (used as a model of human enteropathogenic and enterohaemorrhagic E. coli infections vary between mice strains, from mild self-resolving colonization in NIH Swiss mice to lethality in C3H/HeJ mice. However, no clear genetic component had been shown to be responsible for the differences observed with C. rodentium infections. Because the intestinal microbiota is important in regulating resistance to infection, and microbial composition is dependent on host genotype, it was tested whether variations in microbial composition between mouse strains contributed to differences in "host" susceptibility by transferring the microbiota of resistant mice to lethally susceptible mice prior to infection. Successful transfer of the microbiota from resistant to susceptible mice resulted in delayed pathogen colonization and mortality. Delayed mortality was associated with increased IL-22 mediated innate defense including antimicrobial peptides Reg3γ and Reg3β, and immunono-neutralization of IL-22 abrogated the beneficial effect of microbiota transfer. Conversely, depletion of the native microbiota in resistant mice by antibiotics and transfer of the susceptible mouse microbiota resulted in reduced innate defenses and greater pathology upon infection. This work demonstrates the importance of the microbiota and how it regulates mucosal immunity, providing an important factor in susceptibility to enteric infection. Transfer of resistance through microbial transplantation (bacteriotherapy provides additional mechanisms to alter "host" resistance, and a novel means to alter enteric infection and to study host-pathogen interactions.

  9. U Mineral Hosts and Enrichment Processes in Altered Oceanic Crust

    Science.gov (United States)

    Farr, L. C.; Plank, T.; Kelley, K.; Alt, J. C.

    2001-12-01

    The U-Pb-Th isotopic system is a primary tool for understanding mantle and continental evolution and for quantifying the flow of mass and heat through the Earth's reservoirs. One of the major sites of U-Pb-Th fractionation is the oceanic crust, which is a sink for seawater U. For example, the upper, oxidized oceanic crust (U ~0.4ppm) may be as much as 4x enriched over pristine igneous values (U ~0.09ppm) with a minor net change in Pb and Th. Little, however, is understood about the mechanisms controlling uranium enrichment, its mineral hosts, or the timing of the process. We have used laser ablation inductively coupled mass spectrometry (LA-ICP-MS) and electron microprobe data to study the distribution of U in the oldest sampled crust in the Pacific, Jurassic mid-ocean ridge basalts (MORB) from ODP Site 801C (1000 km seaward of the Mariana trench), formed at fast spreading rates. Seventeen thin sections, 8 with the highest whole rock U content (HUC) (0.61-1.7ppm) and 9 with low U content (LUC) (U laser beam was rastered across various alteration zones, such as halos, veins, and the surrounding host to provide in-situ multi-element analysis (U, Th, Pb, REE, alkalis, etc). HUC are exclusively associated with low-Mg calcites (enrichment, whereas LUC inherit the LREE depletion of the MORB host. Thus, there appear to be distinct generations of fluids that precipitate carbonate; those with the high Sr and low REE of seawater tend to precipitate U-rich calcites (up to 4.5 ppm U). Four thin sections containing Fe-oxide veins ± celadonite and carbonates were also analyzed. Initial analysis suggests high U ( ~1.8ppm) is associated with Fe-oxides, halos immediately surrounding the mixed Fe-oxide veins, or redox fronts further into the basaltic hosts (up to 0.5mm from the oxide veins). Thus, U in altered MORB is associated with three types of alteration phases: 1) carbonate veins, 2) halos surrounding mixed Fe-oxide veins, and 3) Fe-oxide phases. Of the excess U in the upper

  10. Modified host cells with efflux pumps

    Energy Technology Data Exchange (ETDEWEB)

    Dunlop, Mary J.; Keasling, Jay D.; Mukhopadhyay, Aindrila

    2016-08-30

    The present invention provides for a modified host cell comprising a heterologous expression of an efflux pump capable of transporting an organic molecule out of the host cell wherein the organic molecule at a sufficiently high concentration reduces the growth rate of or is lethal to the host cell.

  11. Host coevolution alters the adaptive landscape of a virus

    Science.gov (United States)

    2016-01-01

    The origin of new and complex structures and functions is fundamental for shaping the diversity of life. Such key innovations are rare because they require multiple interacting changes. We sought to understand how the adaptive landscape led to an innovation whereby bacteriophage λ evolved the new ability to exploit a receptor, OmpF, on Escherichia coli cells. Previous work showed that this ability evolved repeatedly, despite requiring four mutations in one virus gene. Here, we examine how this innovation evolved by studying six intermediate genotypes of λ isolated during independent transitions to exploit OmpF and comparing them to their ancestor. All six intermediates showed large increases in their adsorption rates on the ancestral host. Improvements in adsorption were offset, in large part, by the evolution of host resistance, which occurred by reduced expression of LamB, the usual receptor for λ. As a consequence of host coevolution, the adaptive landscape of the virus changed such that selection favouring four of the six virus intermediates became stronger after the host evolved resistance, thereby accelerating virus populations along the path to using the new OmpF receptor. This dependency of viral fitness on host genotype thus shows an important role for coevolution in the origin of the new viral function. PMID:27683370

  12. Exosomes Secreted by Toxoplasma gondii-Infected L6 Cells: Their Effects on Host Cell Proliferation and Cell Cycle Changes

    OpenAIRE

    Kim, Min Jae; Jung, Bong-Kwang; Cho, Jaeeun; Song, Hyemi; Pyo, Kyung-Ho; Lee, Ji Min; Kim, Min-Kyung; Chai, Jong-Yil

    2016-01-01

    Toxoplasma gondii infection induces alteration of the host cell cycle and cell proliferation. These changes are not only seen in directly invaded host cells but also in neighboring cells. We tried to identify whether this alteration can be mediated by exosomes secreted by T. gondii-infected host cells. L6 cells, a rat myoblast cell line, and RH strain of T. gondii were selected for this study. L6 cells were infected with or without T. gondii to isolate exosomes. The cellular growth patterns w...

  13. Salmonella Degrades the Host Glycocalyx Leading to Altered Infection and Glycan Remodeling.

    Science.gov (United States)

    Arabyan, Narine; Park, Dayoung; Foutouhi, Soraya; Weis, Allison M; Huang, Bihua C; Williams, Cynthia C; Desai, Prerak; Shah, Jigna; Jeannotte, Richard; Kong, Nguyet; Lebrilla, Carlito B; Weimer, Bart C

    2016-01-01

    Complex glycans cover the gut epithelial surface to protect the cell from the environment. Invasive pathogens must breach the glycan layer before initiating infection. While glycan degradation is crucial for infection, this process is inadequately understood. Salmonella contains 47 glycosyl hydrolases (GHs) that may degrade the glycan. We hypothesized that keystone genes from the entire GH complement of Salmonella are required to degrade glycans to change infection. This study determined that GHs recognize the terminal monosaccharides (N-acetylneuraminic acid (Neu5Ac), galactose, mannose, and fucose) and significantly (p < 0.05) alter infection. During infection, Salmonella used its two GHs sialidase nanH and amylase malS for internalization by targeting different glycan structures. The host glycans were altered during Salmonella association via the induction of N-glycan biosynthesis pathways leading to modification of host glycans by increasing fucosylation and mannose content, while decreasing sialylation. Gene expression analysis indicated that the host cell responded by regulating more than 50 genes resulting in remodeled glycans in response to Salmonella treatment. This study established the glycan structures on colonic epithelial cells, determined that Salmonella required two keystone GHs for internalization, and left remodeled host glycans as a result of infection. These data indicate that microbial GHs are undiscovered virulence factors. PMID:27389966

  14. Cellular responses to HSV-1 infection are linked to specific types of alterations in the host transcriptome.

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    Hu, Benxia; Li, Xin; Huo, Yongxia; Yu, Yafen; Zhang, Qiuping; Chen, Guijun; Zhang, Yaping; Fraser, Nigel W; Wu, Dongdong; Zhou, Jumin

    2016-01-01

    Pathogen invasion triggers a number of cellular responses and alters the host transcriptome. Here we report that the type of changes to cellular transcriptome is related to the type of cellular functions affected by lytic infection of Herpes Simplex Virus type I in Human primary fibroblasts. Specifically, genes involved in stress responses and nuclear transport exhibited mostly changes in alternative polyadenylation (APA), cell cycle genes showed mostly alternative splicing (AS) changes, while genes in neurogenesis, rarely underwent these changes. Transcriptome wide, the infection resulted in 1,032 cases of AS, 161 incidences of APA, 1,827 events of isoform changes, and up regulation of 596 genes and down regulations of 61 genes compared to uninfected cells. Thus, these findings provided important and specific links between cellular responses to HSV-1 infection and the type of alterations to the host transcriptome, highlighting important roles of RNA processing in virus-host interactions. PMID:27354008

  15. Campylobacter jejuni pdxA affects flagellum-mediated motility to alter host colonization.

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    Hiroshi Asakura

    Full Text Available Vitamin B6 (pyridoxal-5'-phosphate, PLP is linked to a variety of biological functions in prokaryotes. Here, we report that the pdxA (putative 4-hydroxy-L-threonine phosphate dehydrogenase gene plays a pivotal role in the PLP-dependent regulation of flagellar motility, thereby altering host colonization in a leading foodborne pathogen, Campylobacter jejuni. A C. jejuni pdxA mutant failed to produce PLP and exhibited a coincident loss of flagellar motility. Mass spectrometric analyses showed a 3-fold reduction in the main flagellar glycan pseudaminic acid (Pse associated with the disruption of pdxA. The pdxA mutant also exhibited reduced growth rates compared with the WT strain. Comparative metabolomic analyses revealed differences in respiratory/energy metabolism between WT C. jejuni and the pdxA mutant, providing a possible explanation for the differential growth fitness between the two strains. Consistent with the lack of flagellar motility, the pdxA mutant showed impaired motility-mediated responses (bacterial adhesion, ERK1/2 activation, and IL-8 production in INT407 cells and reduced colonization of chickens compared with the WT strain. Overall, this study demonstrated that the pdxA gene affects the PLP-mediated flagellar motility function, mainly through alteration of Pse modification, and the disruption of this gene also alters the respiratory/energy metabolisms to potentially affect host colonization. Our data therefore present novel implications regarding the utility of PLP and its dependent enzymes as potent target(s for the control of this pathogen in the poultry host.

  16. Di-octyl phthalate induced altered host resistance: Viral and protozoal models in mice

    Energy Technology Data Exchange (ETDEWEB)

    Dogra, R.K.S.; Khanna, S.; Srivastava, S.N.; Shukla, L.; Shanker, R. (Industrial Toxicology Research Centre, Lucknow (India)); Chandra, K.; Chandra, S.; Katiyar, J.C. (Central Drug Research Institute, Lucknow (India))

    1989-01-01

    Among industrially important chemicals, the effect of phthalate ester plasticizers on host resistance and immune surveillance to disease has not been well studied. Our recent studies with Di-ccetyl phthalate (DOP) have demonstrated lymphoid organotoxicity, alteration in the functioning of immune system and altered host resistance to a hookworm parasite (Nippostrongylus brasiliensis) in rodents. These observations suggested that DOP, probably through its effect on immune system, could result in altered host resistance to infection. The present studies were, therefore, undertaken to further assess the altered host resistance in DOP treated mice when challenged with either a virus (encephalomyocarditis) or a protozoal (plasmodium) infection, to delineate the possible contribution of phthalate-induced state of immunomodulation to infections.

  17. Epichloe endophytes alter inducible indirect defences in host grasses.

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    Tao Li

    Full Text Available Epichloë endophytes are common symbionts living asymptomatically in pooid grasses and may provide chemical defences against herbivorous insects. While the mechanisms underlying these fungal defences have been well studied, it remains unknown whether endophyte presence affects the host's own defences. We addressed this issue by examining variation in the impact of Epichloë on constitutive and herbivore-induced emissions of volatile organic compounds (VOC, a well-known indirect plant defence, between two grass species, Schedonorus phoenix (ex. Festuca arundinacea; tall fescue and Festuca pratensis (meadow fescue. We found that feeding by a generalist aphid species, Rhopalosiphum padi, induced VOC emissions by uninfected plants of both grass species but to varying extents, while mechanical wounding failed to do so in both species after one day of damage. Interestingly, regardless of damage treatment, Epichloë uncinata-infected F. pratensis emitted significantly lower quantities of VOCs than their uninfected counterparts. In contrast, Epichloë coenophiala-infected S. phoenix did not differ from their uninfected counterparts in constitutive VOC emissions but tended to increase VOC emissions under intense aphid feeding. A multivariate analysis showed that endophyte status imposed stronger differences in VOC profiles of F. pratensis than damage treatment, while the reverse was true for S. phoenix. Additionally, both endophytes inhibited R. padi population growth as measured by aphid dry biomass, with the inhibition appearing greater in E. uncinata-infected F. pratensis. Our results suggest, not only that Epichloë endophytes may play important roles in mediating host VOC responses to herbivory, but also that the magnitude and direction of such responses may vary with the identity of the Epichloë-grass symbiosis. Whether Epichloë-mediated host VOC responses will eventually translate into effects on higher trophic levels merits future investigation.

  18. Exploitation of host cells by Burkholderia pseudomallei.

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    Stevens, Mark P; Galyov, Edouard E

    2004-04-01

    Intracellular bacterial pathogens have evolved mechanisms to enter and exit eukaryotic cells using the power of actin polymerisation and to subvert the activity of cellular enzymes and signal transduction pathways. The proteins deployed by bacteria to subvert cellular processes often mimic eukaryotic proteins in their structure or function. Studies on the exploitation of host cells by the facultative intracellular pathogen Burkholderia pseudomallei are providing novel insights into the pathogenesis of melioidosis, a serious invasive disease of animals and humans that is endemic in tropical and subtropical areas. B. pseudomallei can invade epithelial cells, survive and proliferate inside phagocytes, escape from endocytic vesicles, form actin-based membrane protrusions and induce host cell fusion. Here we review current understanding of the molecular mechanisms underlying these processes.

  19. Alterations of the Host Microbiome Affect Behavioral Responses to Cocaine

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    Kiraly, Drew D.; Walker, Deena M.; Calipari, Erin S.; Labonte, Benoit; Issler, Orna; Pena, Catherine J.; Ribeiro, Efrain A.; Russo, Scott J.; Nestler, Eric J.

    2016-01-01

    Addiction to cocaine and other psychostimulants represents a major public health crisis. The development and persistence of addictive behaviors comes from a complex interaction of genes and environment - the precise mechanisms of which remain elusive. In recent years a surge of evidence has suggested that the gut microbiome can have tremendous impact on behavioral via the microbiota-gut-brain axis. In this study we characterized the influence of the gut microbiota on cocaine-mediated behaviors. Groups of mice were treated with a prolonged course of non-absorbable antibiotics via the drinking water, which resulted in a substantial reduction of gut bacteria. Animals with reduced gut bacteria showed an enhanced sensitivity to cocaine reward and enhanced sensitivity to the locomotor-sensitizing effects of repeated cocaine administration. These behavioral changes were correlated with adaptations in multiple transcripts encoding important synaptic proteins in the brain’s reward circuitry. This study represents the first evidence that alterations in the gut microbiota affect behavioral response to drugs of abuse. PMID:27752130

  20. Phosphorus source alters host plant response to ectomycorrhizal diversity.

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    Baxter, James W; Dighton, John

    2005-11-01

    We examined the influence of phosphorus source and availability on host plant (Pinus rigida) response to ectomycorrhizal diversity under contrasting P conditions. An ectomycorrhizal richness gradient was established with equimolar P supplied as either inorganic phosphate or organic inositol hexaphosphate. We measured growth and N and P uptake of individual P. rigida seedlings inoculated with one, two, or four species of ectomycorrhizal fungi simultaneously and without mycorrhizas in axenic culture. Whereas colonization of P. rigida by individual species of ectomycorrhizal fungi decreased with increasing fungal richness, colonization of all species combined increased. Plant biomass and N content increased across the ectomycorrhizal richness gradient in the organic but not the inorganic P treatment. Plants grown under organic P conditions had higher N concentration than those grown under inorganic P conditions, but there was no effect of richness. Phosphorus content of plants grown in the organic P treatment increased with increasing ectomycorrhizal richness, but there was no response in the inorganic P treatment. Phosphorus concentration was higher in plants grown at the four-species richness level in the organic P treatment, but there was no effect of diversity under inorganic P conditions. Overall, few ectomycorrhizal composition effects were found on plant growth or nutrient status. Phosphatase activities of individual ectomycorrhizal fungi differed under organic P conditions, but there was no difference in total root system phosphatase expression between the inorganic or organic P treatments or across richness levels. Our results provide evidence that plant response to ectomycorrhizal diversity is dependent on the source and availability of P. PMID:15809869

  1. A Systems Survey of Progressive Host-Cell Reorganization during Rotavirus Infection.

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    Green, Victoria A; Pelkmans, Lucas

    2016-07-13

    Pathogen invasion is often accompanied by widespread alterations in cellular physiology, which reflects the hijacking of host factors and processes for pathogen entry and replication. Although genetic perturbation screens have revealed the complexity of host factors involved for numerous pathogens, it has remained challenging to temporally define the progression of events in host cell reorganization during infection. We combine high-confidence genome-scale RNAi screening of host factors required for rotavirus infection in human intestinal cells with an innovative approach to infer the trajectory of virus infection from fixed cell populations. This approach reveals a comprehensive network of host cellular processes involved in rotavirus infection and implicates AMPK in initiating the development of a rotavirus-permissive environment. Our work provides a powerful approach that can be generalized to order complex host cellular requirements along a trajectory of cellular reorganization during pathogen invasion. PMID:27414499

  2. A Systems Survey of Progressive Host-Cell Reorganization during Rotavirus Infection.

    Science.gov (United States)

    Green, Victoria A; Pelkmans, Lucas

    2016-07-13

    Pathogen invasion is often accompanied by widespread alterations in cellular physiology, which reflects the hijacking of host factors and processes for pathogen entry and replication. Although genetic perturbation screens have revealed the complexity of host factors involved for numerous pathogens, it has remained challenging to temporally define the progression of events in host cell reorganization during infection. We combine high-confidence genome-scale RNAi screening of host factors required for rotavirus infection in human intestinal cells with an innovative approach to infer the trajectory of virus infection from fixed cell populations. This approach reveals a comprehensive network of host cellular processes involved in rotavirus infection and implicates AMPK in initiating the development of a rotavirus-permissive environment. Our work provides a powerful approach that can be generalized to order complex host cellular requirements along a trajectory of cellular reorganization during pathogen invasion.

  3. Counting Legionella cells within single amoeba host cells

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    Here we present the first attempt to quantify L. pneumophila cell numbers within individual amoebae hosts that may be released into engineered water systems. The maximum numbers of culturable L. pneumophila cells grown within Acanthamoeba polyphaga and Naegleria fowleri were 134...

  4. Hepatitis C virus host cell interactions uncovered

    DEFF Research Database (Denmark)

    Gottwein, Judith; Bukh, Jens

    2007-01-01

      Insights into virus-host cell interactions as uncovered by Randall et al. (1) in a recent issue of PNAS further our understanding of the hepatitis C virus (HCV) life cycle, persistence, and pathogenesis and might lead to the identification of new therapeutic targets. HCV persistently infects 180...... treated. Therefore, there is a pressing need for the identification of novel drugs against hepatitis C. Although most research focuses on the development of HCV-specific antivirals, such as protease and polymerase inhibitors (3), cellular targets could be pursued and might allow the development of broad...

  5. Novel chikungunya vaccine candidate with an IRES-based attenuation and host range alteration mechanism.

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    Kenneth Plante

    2011-07-01

    Full Text Available Chikungunya virus (CHIKV is a reemerging mosquito-borne pathogen that has recently caused devastating urban epidemics of severe and sometimes chronic arthralgia. As with most other mosquito-borne viral diseases, control relies on reducing mosquito populations and their contact with people, which has been ineffective in most locations. Therefore, vaccines remain the best strategy to prevent most vector-borne diseases. Ideally, vaccines for diseases of resource-limited countries should combine low cost and single dose efficacy, yet induce rapid and long-lived immunity with negligible risk of serious adverse reactions. To develop such a vaccine to protect against chikungunya fever, we employed a rational attenuation mechanism that also prevents the infection of mosquito vectors. The internal ribosome entry site (IRES from encephalomyocarditis virus replaced the subgenomic promoter in a cDNA CHIKV clone, thus altering the levels and host-specific mechanism of structural protein gene expression. Testing in both normal outbred and interferon response-defective mice indicated that the new vaccine candidate is highly attenuated, immunogenic and efficacious after a single dose. Furthermore, it is incapable of replicating in mosquito cells or infecting mosquitoes in vivo. This IRES-based attenuation platform technology may be useful for the predictable attenuation of any alphavirus.

  6. Differential proteome analysis of chikungunya virus infection on host cells.

    Directory of Open Access Journals (Sweden)

    Christina Li-Ping Thio

    Full Text Available BACKGROUND: Chikungunya virus (CHIKV is an emerging mosquito-borne alphavirus that has caused multiple unprecedented and re-emerging outbreaks in both tropical and temperate countries. Despite ongoing research efforts, the underlying factors involved in facilitating CHIKV replication during early infection remains ill-characterized. The present study serves to identify host proteins modulated in response to early CHIKV infection using a proteomics approach. METHODOLOGY AND PRINCIPAL FINDINGS: The whole cell proteome profiles of CHIKV-infected and mock control WRL-68 cells were compared and analyzed using two-dimensional gel electrophoresis (2-DGE. Fifty-three spots were found to be differentially modulated and 50 were successfully identified by MALDI-TOF/TOF. Eight were significantly up-regulated and 42 were down-regulated. The mRNA expressions of 15 genes were also found to correlate with the corresponding protein expression. STRING network analysis identified several biological processes to be affected, including mRNA processing, translation, energy production and cellular metabolism, ubiquitin-proteasome pathway (UPP and cell cycle regulation. CONCLUSION/SIGNIFICANCE: This study constitutes a first attempt to investigate alteration of the host cellular proteome during early CHIKV infection. Our proteomics data showed that during early infection, CHIKV affected the expression of proteins that are involved in mRNA processing, host metabolic machinery, UPP, and cyclin-dependent kinase 1 (CDK1 regulation (in favour of virus survival, replication and transmission. While results from this study complement the proteomics results obtained from previous late host response studies, functional characterization of these proteins is warranted to reinforce our understanding of their roles during early CHIKV infection in humans.

  7. Cement degradation and the alteration of host rocks. Studies within the Grimsel Test Site Project.

    Science.gov (United States)

    Soler, J. M.

    2009-04-01

    Cement is a major component of the engineered barrier system in proposed underground repositories for low- and intermediate-level radioactive waste. Cement grouting of highly-conductive fractures in the vicinity of such repositories is also planned. The interaction between the hyperalkaline solutions derived from the degradation of cement and the rocks hosting such repositories may change the physical and chemical properties of the host rocks. The HPF project (Hyperalkaline Plume in Fractured Rock; ANDRA-FR-, DOE-USA-, JAEA-JP-, NAGRA-CH-, POSIVA-FI-, SKB-SE-) studied the alteration of a fractured granite due to the circulation of a synthetic high-pH solution. A significant decrease in fracture permeability was observed both in the laboratory (core infiltration experiment; decimeter scale) and in the Grimsel Test Site (circulation along a fracture; meter scale), despite the relatively minor mineralogical alteration. Coupling of mineralogical alteration and permeability changes was incorporated into reactive transport modeling of the experiments. The hydration and degradation of cement are being explicitly incorporated into the new LCS (Long-Term Cement Studies; JAEA-JP-, NAGRA-CH-, NDA-GB-, POSIVA-FI-) project at Grimsel. New laboratory and field experiments including a cement source are being designed. Reactive transport modeling of the degradation of cement, causing the formation of hyperalkaline solutions and the alteration of the host rock, will be an essential part of the experiment.

  8. Acanthamoeba induces cell-cycle arrest in host cells

    OpenAIRE

    Sissons, J.; Alsam, S.; Jayasekera, S.; Kim, K S; Stins, M; Khan, Naveed Ahmed

    2004-01-01

    Acanthamoeba can cause fatal granulomatous amoebic encephalitis (GAE) and eye keratitis. However, the pathogenesis and pathophysiology of these emerging diseases remain unclear. In this study, the effects of Acanthamoeba on the host cell cycle using human brain microvascular endothelial cells (HBMEC) and human corneal epithelial cells (HCEC) were determined. Two isolates of Acanthamoeba belonging to the T1 genotype (GAE isolate) and T4 genotype (keratitis isolate) were used, which showed seve...

  9. Climate change and infectious diseases of wildlife: Altered interactions between pathogens, vectors and hosts

    Directory of Open Access Journals (Sweden)

    Milena GALLANA, Marie-Pierre RYSER-DEGIORGIS, Thomas WAHLI, Helmut SEGNER

    2013-06-01

    Full Text Available Infectious diseases result from the interactions of host, pathogens, and, in the case of vector-borne diseases, also vectors. The interactions involve physiological and ecological mechanisms and they have evolved under a given set of environmental conditions. Environmental change, therefore, will alter host-pathogen-vector interactions and, consequently, the distribution, intensity, and dynamics of infectious diseases. Here, we review how climate change may impact infectious diseases of aquatic and terrestrial wildlife. Climate change can have direct impacts on distribution, life cycle, and physiological status of hosts, pathogens and vectors. While a change in either host, pathogen or vector does not necessarily translate into an alteration of the disease, it is the impact of climate change on the interactions between the disease components which is particularly critical for altered disease risks. Finally, climate factors can modulate disease through modifying the ecological networks host-pathogen-vector systems are belonging to, and climate change can combine with other environmental stressors to induce cumulative effects on infectious diseases. Overall, the influence of climate change on infectious diseases involves different mechanisms, it can be modulated by phenotypic acclimation and/or genotypic adaptation, it depends on the ecological context of the host-pathogen-vector interactions, and it can be modulated by impacts of other stressors. As a consequence of this complexity, non-linear responses of disease systems under climate change are to be expected. To improve predictions on climate change impacts on infectious disease, we suggest that more emphasis should be given to the integration of biomedical and ecological research for studying both the physiological and ecological mechanisms which mediate climate change impacts on disease, and to the development of harmonized methods and approaches to obtain more comparable results, as this

  10. Climate change and infectious diseases of wildlife: Altered interactions between pathogens, vectors and hosts

    Institute of Scientific and Technical Information of China (English)

    Milena GALLANA; Marie-Pierre RYSER-DEGIORGIS; Thomas WAHLI; Helmut SEGNER

    2013-01-01

    Infectious diseases result from the interactions of host,pathogens,and,in the case of vector-borne diseases,also vectors.The interactions involve physiological and ecological mechanisms and they have evolved under a given set of environmental conditions.Environmental change,therefore,will alter host-pathogen-vector interactions and,consequently,the distribution,intensity,and dynamics of infectious diseases.Here,we review how climate change may impact infectious diseases of aquatic and terrestrial wildlife.Climate change can have direct impacts on distribution,life cycle,and physiological status of hosts,pathogens and vectors.While a change in either host,pathogen or vector does not necessarily translate into an alteration of the disease,it is the impact of climate change on the interactions between the disease components which is particularly critical for altered disease risks.Finally,climate factors can modulate disease through modifying the ecological networks host-pathogen-vector systems are belonging to,and climate change can combine with other environmental stressors to induce cumulative effects on infectious diseases.Overall,the influence of climate change on infectious diseases involves different mechanisms,it can be modulated by phenotypic acclimation and/or genotypic adaptation,it depends on the ecological context of the host-pathogen-vector interactions,and it can be modulated by impacts of other stressors.As a consequence of this complexity,non-linear responses of disease systems under climate change are to be expected.To improve predictions on climate change impacts on infectious disease,we suggest that more emphasis should be given to the integration of biomedical and ecological research for studying both the physiological and ecological mechanisms which mediate climate change impacts on disease,and to the development of harmonized methods and approaches to obtain more comparable results,as this would support the discrimination of case-specific versus

  11. Toxocara in the mouse : A model for parasite-altered host behaviour ?

    OpenAIRE

    Holland, Celia

    2001-01-01

    The objective of this paper is to critically evaluate the significance of parasite-altered host behaviour in the Toxocara mouse model particularly in the light of the Manipulation Hypothesis. Murine behaviours were examined in both outbred and inbred strains of mice infected with different doses of Toxocara canis ova. Behaviours investigated included activity, exploration, response to novelty, anxiety, learning, memory and social behaviour. Subsequent modifications to the behaviour of infecte...

  12. Understanding and altering cell tropism of vesicular stomatitis virus

    Science.gov (United States)

    Hastie, Eric; Cataldi, Marcela; Marriott, Ian; Grdzelishvili, Valery Z.

    2013-01-01

    Vesicular stomatitis virus (VSV) is a prototypic nonsegmented negative-strand RNA virus. VSV’s broad cell tropism makes it a popular model virus for many basic research applications. In addition, a lack of preexisting human immunity against VSV, inherent oncotropism and other features make VSV a widely used platform for vaccine and oncolytic vectors. However, VSV’s neurotropism that can result in viral encephalitis in experimental animals needs to be addressed for the use of the virus as a safe vector. Therefore, it is very important to understand the determinants of VSV tropism and develop strategies to alter it. VSV glycoprotein (G) and matrix (M) protein play major roles in its cell tropism. VSV G protein is responsible for VSV broad cell tropism and is often used for pseudotyping other viruses. VSV M affects cell tropism via evasion of antiviral responses, and M mutants can be used to limit cell tropism to cell types defective in interferon signaling. In addition, other VSV proteins and host proteins may function as determinants of VSV cell tropism. Various approaches have been successfully used to alter VSV tropism to benefit basic research and clinically relevant applications. PMID:23796410

  13. Host-based Th2 cell therapy for prolongation of cardiac allograft viability.

    Directory of Open Access Journals (Sweden)

    Shoba Amarnath

    Full Text Available Donor T cell transfusion, which is a long-standing approach to prevent allograft rejection, operates indirectly by alteration of host T cell immunity. We therefore hypothesized that adoptive transfer of immune regulatory host Th2 cells would represent a novel intervention to enhance cardiac allograft survival. Using a well-described rat cardiac transplant model, we first developed a method for ex vivo manufacture of rat host-type Th2 cells in rapamycin, with subsequent injection of such Th2.R cells prior to class I and class II disparate cardiac allografting. Second, we determined whether Th2.R cell transfer polarized host immunity towards a Th2 phenotype. And third, we evaluated whether Th2.R cell therapy prolonged allograft viability when used alone or in combination with a short-course of cyclosporine (CSA therapy. We found that host-type Th2.R cell therapy prior to cardiac allografting: (1 reduced the frequency of activated T cells in secondary lymphoid organs; (2 shifted post-transplant cytokines towards a Th2 phenotype; and (3 prolonged allograft viability when used in combination with short-course CSA therapy. These results provide further support for the rationale to use "direct" host T cell therapy for prolongation of allograft viability as an alternative to "indirect" therapy mediated by donor T cell infusion.

  14. A host cell membrane microdomain is a critical factor for organelle discharge by Toxoplasma gondii.

    Science.gov (United States)

    Tahara, Michiru; Andrabi, Syed Bilal Ahmad; Matsubara, Ryuma; Aonuma, Hiroka; Nagamune, Kisaburo

    2016-10-01

    Host cell microdomains are involved in the attachment, entry, and replication of intracellular microbial pathogens. Entry into the host cell of Toxoplasma gondii and the subsequent survival of this protozoan parasite are tightly coupled with the proteins secreted from organelle called rhoptry. The rhoptry proteins are rapidly discharged into clusters of vesicles, called evacuoles, which are then delivered to parasitophorous vacuoles (PVs) or nucleus. In this study, we examined the roles of two host cell microdomain components, cholesterol and glycosylphosphatidylinositol (GPI), in evacuole formation. The acute depletion of cholesterol from the host cell plasma membrane blocked evacuole formation but not invasion. Whereas the lack of host cell GPI also altered evacuole formation but not invasion, instead inducing excess evacuole formation. The latter effect was not influenced by the evacuole-inhibiting effects of host cell cholesterol depletion, indicating the independent roles of host GPI and cholesterol in evacuole formation. In addition, the excess formation of evacuoles resulted in the enhanced recruitment of host mitochondria and endoplasmic reticulum to PVs, which in turn stimulated the growth of the parasite. PMID:27217289

  15. Radiofrequency treatment alters cancer cell phenotype

    Science.gov (United States)

    Ware, Matthew J.; Tinger, Sophia; Colbert, Kevin L.; Corr, Stuart J.; Rees, Paul; Koshkina, Nadezhda; Curley, Steven; Summers, H. D.; Godin, Biana

    2015-07-01

    The importance of evaluating physical cues in cancer research is gradually being realized. Assessment of cancer cell physical appearance, or phenotype, may provide information on changes in cellular behavior, including migratory or communicative changes. These characteristics are intrinsically different between malignant and non-malignant cells and change in response to therapy or in the progression of the disease. Here, we report that pancreatic cancer cell phenotype was altered in response to a physical method for cancer therapy, a non-invasive radiofrequency (RF) treatment, which is currently being developed for human trials. We provide a battery of tests to explore these phenotype characteristics. Our data show that cell topography, morphology, motility, adhesion and division change as a result of the treatment. These may have consequences for tissue architecture, for diffusion of anti-cancer therapeutics and cancer cell susceptibility within the tumor. Clear phenotypical differences were observed between cancerous and normal cells in both their untreated states and in their response to RF therapy. We also report, for the first time, a transfer of microsized particles through tunneling nanotubes, which were produced by cancer cells in response to RF therapy. Additionally, we provide evidence that various sub-populations of cancer cells heterogeneously respond to RF treatment.

  16. Metabolic alterations in renal cell carcinoma.

    Science.gov (United States)

    Massari, Francesco; Ciccarese, Chiara; Santoni, Matteo; Brunelli, Matteo; Piva, Francesco; Modena, Alessandra; Bimbatti, Davide; Fantinel, Emanuela; Santini, Daniele; Cheng, Liang; Cascinu, Stefano; Montironi, Rodolfo; Tortora, Giampaolo

    2015-11-01

    Renal cell carcinoma (RCC) is a metabolic disease, being characterized by the dysregulation of metabolic pathways involved in oxygen sensing (VHL/HIF pathway alterations and the subsequent up-regulation of HIF-responsive genes such as VEGF, PDGF, EGF, and glucose transporters GLUT1 and GLUT4, which justify the RCC reliance on aerobic glycolysis), energy sensing (fumarate hydratase-deficient, succinate dehydrogenase-deficient RCC, mutations of HGF/MET pathway resulting in the metabolic Warburg shift marked by RCC increased dependence on aerobic glycolysis and the pentose phosphate shunt, augmented lipogenesis, and reduced AMPK and Krebs cycle activity) and/or nutrient sensing cascade (deregulation of AMPK-TSC1/2-mTOR and PI3K-Akt-mTOR pathways). We analyzed the key metabolic abnormalities underlying RCC carcinogenesis, highlighting those altered pathways that may represent potential targets for the development of more effective therapeutic strategies.

  17. Temporal expression of bacterial proteins instructs host CD4 T cell expansion and Th17 development.

    Directory of Open Access Journals (Sweden)

    Seung-Joo Lee

    2012-01-01

    Full Text Available Pathogens can substantially alter gene expression within an infected host depending on metabolic or virulence requirements in different tissues, however, the effect of these alterations on host immunity are unclear. Here we visualized multiple CD4 T cell responses to temporally expressed proteins in Salmonella-infected mice. Flagellin-specific CD4 T cells expanded and contracted early, differentiated into Th1 and Th17 lineages, and were enriched in mucosal tissues after oral infection. In contrast, CD4 T cells responding to Salmonella Type-III Secretion System (TTSS effectors steadily accumulated until bacterial clearance was achieved, primarily differentiated into Th1 cells, and were predominantly detected in systemic tissues. Thus, pathogen regulation of antigen expression plays a major role in orchestrating the expansion, differentiation, and location of antigen-specific CD4 T cells in vivo.

  18. Host cells and methods for production of isobutanol

    Energy Technology Data Exchange (ETDEWEB)

    Anthony, Larry Cameron; He, Hongxian; Huang, Lixuan Lisa; Okeefe, Daniel P.; Kruckeberg, Arthur Leo; Li, Yougen; Maggio-Hall, Lori Ann; McElvain, Jessica; Nelson, Mark J.; Patnaik, Ranjan; Rothman, Steven Cary

    2016-08-23

    Provided herein are recombinant yeast host cells and methods for their use for production of isobutanol. Yeast host cells provided comprise an isobutanol biosynthetic pathway and at least one of reduced or eliminated aldehyde dehydrogenase activity, reduced or eliminated acetolactate reductase activity; or a heterologous polynucleotide encoding a polypeptide having ketol-acid reductoisomerase activity.

  19. How pathogens use linear motifs to perturb host cell networks

    KAUST Repository

    Via, Allegra

    2015-01-01

    Molecular mimicry is one of the powerful stratagems that pathogens employ to colonise their hosts and take advantage of host cell functions to guarantee their replication and dissemination. In particular, several viruses have evolved the ability to interact with host cell components through protein short linear motifs (SLiMs) that mimic host SLiMs, thus facilitating their internalisation and the manipulation of a wide range of cellular networks. Here we present convincing evidence from the literature that motif mimicry also represents an effective, widespread hijacking strategy in prokaryotic and eukaryotic parasites. Further insights into host motif mimicry would be of great help in the elucidation of the molecular mechanisms behind host cell invasion and the development of anti-infective therapeutic strategies.

  20. Streptococcus pneumoniae Infection of Host Epithelial Cells via Polymeric Immunoglobulin Receptor Transiently Induces Calcium Release from Intracellular Stores*

    OpenAIRE

    Asmat, T. M.; Agarwal, V; Rath, S.; Hildebrandt, J.-P.; Hammerschmidt, S.

    2011-01-01

    The pneumococcal surface protein C (PspC) is a major adhesin of Streptococcus pneumoniae (pneumococci) that interacts in a human-specific manner with the ectodomain of the human polymeric immunoglobulin receptor (pIgR) produced by respiratory epithelial cells. This interaction promotes bacterial colonization and bacterial internalization by initiating host signal transduction cascades. Here, we examined alterations of intracellular calcium ([Ca2+]i) levels in epithelial cells during host cell...

  1. Altered calcium signaling in cancer cells.

    Science.gov (United States)

    Stewart, Teneale A; Yapa, Kunsala T D S; Monteith, Gregory R

    2015-10-01

    It is the nature of the calcium signal, as determined by the coordinated activity of a suite of calcium channels, pumps, exchangers and binding proteins that ultimately guides a cell's fate. Deregulation of the calcium signal is often deleterious and has been linked to each of the 'cancer hallmarks'. Despite this, we do not yet have a full understanding of the remodeling of the calcium signal associated with cancer. Such an understanding could aid in guiding the development of therapies specifically targeting altered calcium signaling in cancer cells during tumorigenic progression. Findings from some of the studies that have assessed the remodeling of the calcium signal associated with tumorigenesis and/or processes important in invasion and metastasis are presented in this review. The potential of new methodologies is also discussed. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers.

  2. Inhibition of host cell translation elongation by Legionella pneumophila blocks the host cell unfolded protein response

    Science.gov (United States)

    Hempstead, Andrew D.; Isberg, Ralph R.

    2015-01-01

    Cells of the innate immune system recognize bacterial pathogens by detecting common microbial patterns as well as pathogen-specific activities. One system that responds to these stimuli is the IRE1 branch of the unfolded protein response (UPR), a sensor of endoplasmic reticulum (ER) stress. Activation of IRE1, in the context of Toll-like receptor (TLR) signaling, induces strong proinflammatory cytokine induction. We show here that Legionella pneumophila, an intravacuolar pathogen that replicates in an ER-associated compartment, blocks activation of the IRE1 pathway despite presenting pathogen products that stimulate this response. L. pneumophila TLR ligands induced the splicing of mRNA encoding XBP1s, the main target of IRE1 activity. L. pneumophila was able to inhibit both chemical and bacterial induction of XBP1 splicing via bacterial translocated proteins that interfere with host protein translation. A strain lacking five translocated translation elongation inhibitors was unable to block XBP1 splicing, but this could be rescued by expression of a single such inhibitor, consistent with limitation of the response by translation elongation inhibitors. Chemical inhibition of translation elongation blocked pattern recognition receptor-mediated XBP1 splicing, mimicking the effects of the bacterial translation inhibitors. In contrast, host cell-promoted inhibition of translation initiation in response to the pathogen was ineffective in blocking XBP1 splicing, demonstrating the need for the elongation inhibitors for protection from the UPR. The inhibition of host translation elongation may be a common strategy used by pathogens to limit the innate immune response by interfering with signaling via the UPR. PMID:26598709

  3. Control of host cell phosphorylation by Legionella pneumophila

    Directory of Open Access Journals (Sweden)

    Eva eHaenssler

    2011-04-01

    Full Text Available Phosphorylation is one of the most frequent modifications in intracellular signaling and is implicated in many processes ranging from transcriptional control to signal transduction in innate immunity. Many pathogens modulate host cell phosphorylation pathways to promote growth and establish an infectious disease. The intracellular pathogen L. pneumophila targets and exploits the host phosphorylation system throughout the infection cycle as part of its strategy to establish an environment beneficial for replication. Key to this manipulation is the L. pneumophila Icm/Dot type IV secretion system, which translocates bacterial proteins into the host cytosol that can act directly on phosphorylation cascades. This review will focus on the different stages of L. pneumophila infection, in which host kinases and phosphatases contribute to infection of the host cell and promote intracellular survival of the pathogen. This includes the involvement of phosphatidylinositol 3-kinases during phagocytosis as well as the role of phosphoinositide metabolism during the establishment of the replication vacuole. Furthermore, L. pneumophila infection modulates the NF-κB and MAPK pathways, two signaling pathways that are central to the host innate immune response and involved in regulation of host cell survival. Therefore, L. pneumophila infection manipulates host cell signal transduction by phosphorylation at multiple levels.

  4. Virus Infection of Plants Alters Pollinator Preference: A Payback for Susceptible Hosts?

    Science.gov (United States)

    Groen, Simon C; Jiang, Sanjie; Murphy, Alex M; Cunniffe, Nik J; Westwood, Jack H; Davey, Matthew P; Bruce, Toby J A; Caulfield, John C; Furzer, Oliver J; Reed, Alison; Robinson, Sophie I; Miller, Elizabeth; Davis, Christopher N; Pickett, John A; Whitney, Heather M; Glover, Beverley J; Carr, John P

    2016-08-01

    Plant volatiles play important roles in attraction of certain pollinators and in host location by herbivorous insects. Virus infection induces changes in plant volatile emission profiles, and this can make plants more attractive to insect herbivores, such as aphids, that act as viral vectors. However, it is unknown if virus-induced alterations in volatile production affect plant-pollinator interactions. We found that volatiles emitted by cucumber mosaic virus (CMV)-infected tomato (Solanum lycopersicum) and Arabidopsis thaliana plants altered the foraging behaviour of bumblebees (Bombus terrestris). Virus-induced quantitative and qualitative changes in blends of volatile organic compounds emitted by tomato plants were identified by gas chromatography-coupled mass spectrometry. Experiments with a CMV mutant unable to express the 2b RNA silencing suppressor protein and with Arabidopsis silencing mutants implicate microRNAs in regulating emission of pollinator-perceivable volatiles. In tomato, CMV infection made plants emit volatiles attractive to bumblebees. Bumblebees pollinate tomato by 'buzzing' (sonicating) the flowers, which releases pollen and enhances self-fertilization and seed production as well as pollen export. Without buzz-pollination, CMV infection decreased seed yield, but when flowers of mock-inoculated and CMV-infected plants were buzz-pollinated, the increased seed yield for CMV-infected plants was similar to that for mock-inoculated plants. Increased pollinator preference can potentially increase plant reproductive success in two ways: i) as female parents, by increasing the probability that ovules are fertilized; ii) as male parents, by increasing pollen export. Mathematical modeling suggested that over a wide range of conditions in the wild, these increases to the number of offspring of infected susceptible plants resulting from increased pollinator preference could outweigh underlying strong selection pressures favoring pathogen resistance

  5. Virus Infection of Plants Alters Pollinator Preference: A Payback for Susceptible Hosts?

    Science.gov (United States)

    Davey, Matthew P.; Bruce, Toby J. A.; Caulfield, John C.; Furzer, Oliver J.; Reed, Alison; Robinson, Sophie I.; Miller, Elizabeth; Davis, Christopher N.; Pickett, John A.; Whitney, Heather M.; Glover, Beverley J.; Carr, John P.

    2016-01-01

    Plant volatiles play important roles in attraction of certain pollinators and in host location by herbivorous insects. Virus infection induces changes in plant volatile emission profiles, and this can make plants more attractive to insect herbivores, such as aphids, that act as viral vectors. However, it is unknown if virus-induced alterations in volatile production affect plant-pollinator interactions. We found that volatiles emitted by cucumber mosaic virus (CMV)-infected tomato (Solanum lycopersicum) and Arabidopsis thaliana plants altered the foraging behaviour of bumblebees (Bombus terrestris). Virus-induced quantitative and qualitative changes in blends of volatile organic compounds emitted by tomato plants were identified by gas chromatography-coupled mass spectrometry. Experiments with a CMV mutant unable to express the 2b RNA silencing suppressor protein and with Arabidopsis silencing mutants implicate microRNAs in regulating emission of pollinator-perceivable volatiles. In tomato, CMV infection made plants emit volatiles attractive to bumblebees. Bumblebees pollinate tomato by ‘buzzing’ (sonicating) the flowers, which releases pollen and enhances self-fertilization and seed production as well as pollen export. Without buzz-pollination, CMV infection decreased seed yield, but when flowers of mock-inoculated and CMV-infected plants were buzz-pollinated, the increased seed yield for CMV-infected plants was similar to that for mock-inoculated plants. Increased pollinator preference can potentially increase plant reproductive success in two ways: i) as female parents, by increasing the probability that ovules are fertilized; ii) as male parents, by increasing pollen export. Mathematical modeling suggested that over a wide range of conditions in the wild, these increases to the number of offspring of infected susceptible plants resulting from increased pollinator preference could outweigh underlying strong selection pressures favoring pathogen resistance

  6. Virus Infection of Plants Alters Pollinator Preference: A Payback for Susceptible Hosts?

    Science.gov (United States)

    Groen, Simon C; Jiang, Sanjie; Murphy, Alex M; Cunniffe, Nik J; Westwood, Jack H; Davey, Matthew P; Bruce, Toby J A; Caulfield, John C; Furzer, Oliver J; Reed, Alison; Robinson, Sophie I; Miller, Elizabeth; Davis, Christopher N; Pickett, John A; Whitney, Heather M; Glover, Beverley J; Carr, John P

    2016-08-01

    Plant volatiles play important roles in attraction of certain pollinators and in host location by herbivorous insects. Virus infection induces changes in plant volatile emission profiles, and this can make plants more attractive to insect herbivores, such as aphids, that act as viral vectors. However, it is unknown if virus-induced alterations in volatile production affect plant-pollinator interactions. We found that volatiles emitted by cucumber mosaic virus (CMV)-infected tomato (Solanum lycopersicum) and Arabidopsis thaliana plants altered the foraging behaviour of bumblebees (Bombus terrestris). Virus-induced quantitative and qualitative changes in blends of volatile organic compounds emitted by tomato plants were identified by gas chromatography-coupled mass spectrometry. Experiments with a CMV mutant unable to express the 2b RNA silencing suppressor protein and with Arabidopsis silencing mutants implicate microRNAs in regulating emission of pollinator-perceivable volatiles. In tomato, CMV infection made plants emit volatiles attractive to bumblebees. Bumblebees pollinate tomato by 'buzzing' (sonicating) the flowers, which releases pollen and enhances self-fertilization and seed production as well as pollen export. Without buzz-pollination, CMV infection decreased seed yield, but when flowers of mock-inoculated and CMV-infected plants were buzz-pollinated, the increased seed yield for CMV-infected plants was similar to that for mock-inoculated plants. Increased pollinator preference can potentially increase plant reproductive success in two ways: i) as female parents, by increasing the probability that ovules are fertilized; ii) as male parents, by increasing pollen export. Mathematical modeling suggested that over a wide range of conditions in the wild, these increases to the number of offspring of infected susceptible plants resulting from increased pollinator preference could outweigh underlying strong selection pressures favoring pathogen resistance

  7. Fungal invasion of normally non-phagocytic host cells.

    Directory of Open Access Journals (Sweden)

    Scott G Filler

    2006-12-01

    Full Text Available Many fungi that cause invasive disease invade host epithelial cells during mucosal and respiratory infection, and subsequently invade endothelial cells during hematogenous infection. Most fungi invade these normally non-phagocytic host cells by inducing their own uptake. Candida albicans hyphae interact with endothelial cells in vitro by binding to N-cadherin on the endothelial cell surface. This binding induces rearrangement of endothelial cell microfilaments, which results in the endocytosis of the organism. The capsule of Cryptococcus neoformans is composed of glucuronoxylomannan, which binds specifically to brain endothelial cells, and appears to mediate both adherence and induction of endocytosis. The mechanisms by which other fungal pathogens induce their own uptake are largely unknown. Some angioinvasive fungi, such as Aspergillus species and the Zygomycetes, invade endothelial cells from the abluminal surface during the initiation of invasive disease, and subsequently invade the luminal surface of endothelial cells during hematogenous dissemination. Invasion of normally non-phagocytic host cells has different consequences, depending on the type of invading fungus. Aspergillus fumigatus blocks apoptosis of pulmonary epithelial cells, whereas Paracoccidioides brasiliensis induces apoptosis of epithelial cells. This review summarizes the mechanisms by which diverse fungal pathogens invade normally non-phagocytic host cells and discusses gaps in our knowledge that provide opportunities for future research.

  8. Cell proliferation alterations in Chlorella cells under stress conditions

    International Nuclear Information System (INIS)

    Very little is known about growth and proliferation in relation to the cell cycle regulation of algae. The lack of knowledge is even greater when referring to the potential toxic effects of pollutants on microalgal cell division. To assess the effect of terbutryn, a triazine herbicide, on the proliferation of the freshwater microalga Chlorella vulgaris three flow cytometric approaches were used: (1) in vivo cell division using 5-,6-carboxyfluorescein diacetate succinimidyl ester (CFSE) staining was measured, (2) the growth kinetics were determined by cytometric cell counting and (3) cell viability was evaluated with the membrane-impermeable double-stranded nucleic acid stain propidium iodide (PI). The results obtained in the growth kinetics study using CFSE to identify the microalgal cell progeny were consistent with those determined by cytometric cell counting. In all C. vulgaris cultures, each mother cell had undergone only one round of division through the 96 h of assay and the cell division occurred during the dark period. Cell division of the cultures exposed to the herbicide was asynchronous. Terbutryn altered the normal number of daughter cells (4 autospores) obtained from each mother cell. The number was only two in the cultures treated with 250 nM. The duration of the lag phase after the exposure to terbutryn could be dependent on the existence of a critical cell size to activate cytoplasmic division. Cell size, complexity and fluorescence of chlorophyll a of the microalgal cells presented a marked light/dark (day/night) cycle, except in the non-dividing 500 nM cultures, where terbutryn arrested cell division at the beginning of the cycle. Viability results showed that terbutryn has an algastatic effect in C. vulgaris cells at this concentration. The rapid and precise determination of cell proliferation by CFSE staining has allowed us to develop a model for assessing both the cell cycle of C. vulgaris and the in vivo effects of pollutants on growth and

  9. Cell proliferation alterations in Chlorella cells under stress conditions

    Energy Technology Data Exchange (ETDEWEB)

    Rioboo, Carmen [Laboratorio de Microbiologia, Facultad de Ciencias, Universidad de A Coruna, Campus da Zapateira s/n, 15008 A Coruna (Spain); O' Connor, Jose Enrique [Laboratorio de Citomica, Unidad Mixta de Investigacion CIPF-UVEG, Centro de Investigacion Principe Felipe, Avda. Autopista del Saler, 16, 46013 Valencia (Spain); Prado, Raquel; Herrero, Concepcion [Laboratorio de Microbiologia, Facultad de Ciencias, Universidad de A Coruna, Campus da Zapateira s/n, 15008 A Coruna (Spain); Cid, Angeles, E-mail: cid@udc.es [Laboratorio de Microbiologia, Facultad de Ciencias, Universidad de A Coruna, Campus da Zapateira s/n, 15008 A Coruna (Spain)

    2009-09-14

    Very little is known about growth and proliferation in relation to the cell cycle regulation of algae. The lack of knowledge is even greater when referring to the potential toxic effects of pollutants on microalgal cell division. To assess the effect of terbutryn, a triazine herbicide, on the proliferation of the freshwater microalga Chlorella vulgaris three flow cytometric approaches were used: (1) in vivo cell division using 5-,6-carboxyfluorescein diacetate succinimidyl ester (CFSE) staining was measured, (2) the growth kinetics were determined by cytometric cell counting and (3) cell viability was evaluated with the membrane-impermeable double-stranded nucleic acid stain propidium iodide (PI). The results obtained in the growth kinetics study using CFSE to identify the microalgal cell progeny were consistent with those determined by cytometric cell counting. In all C. vulgaris cultures, each mother cell had undergone only one round of division through the 96 h of assay and the cell division occurred during the dark period. Cell division of the cultures exposed to the herbicide was asynchronous. Terbutryn altered the normal number of daughter cells (4 autospores) obtained from each mother cell. The number was only two in the cultures treated with 250 nM. The duration of the lag phase after the exposure to terbutryn could be dependent on the existence of a critical cell size to activate cytoplasmic division. Cell size, complexity and fluorescence of chlorophyll a of the microalgal cells presented a marked light/dark (day/night) cycle, except in the non-dividing 500 nM cultures, where terbutryn arrested cell division at the beginning of the cycle. Viability results showed that terbutryn has an algastatic effect in C. vulgaris cells at this concentration. The rapid and precise determination of cell proliferation by CFSE staining has allowed us to develop a model for assessing both the cell cycle of C. vulgaris and the in vivo effects of pollutants on growth and

  10. A mutualistic endophyte alters the niche dimensions of its host plant.

    Science.gov (United States)

    Kazenel, Melanie R; Debban, Catherine L; Ranelli, Luciana; Hendricks, Will Q; Chung, Y Anny; Pendergast, Thomas H; Charlton, Nikki D; Young, Carolyn A; Rudgers, Jennifer A

    2015-01-01

    Mutualisms can play important roles in influencing species coexistence and determining community composition. However, few studies have tested whether such interactions can affect species distributions by altering the niches of partner species. In subalpine meadows of the Rocky Mountains, USA, we explored whether the presence of a fungal endophyte (genus Epichloë) may shift the niche of its partner plant, marsh bluegrass (Poa leptocoma) relative to a closely related but endophyte-free grass species, nodding bluegrass (Poa reflexa). Using observations and a 3-year field experiment, we tested two questions: (i) Do P. leptocoma and P. reflexa occupy different ecological niches? and (ii) Does endophyte presence affect the relative fitness of P. leptocoma versus P. reflexa in the putative niches of these grass species? The two species were less likely to co-occur than expected by chance. Specifically, P. leptocoma grew closer to water sources and in wetter soils than P. reflexa, and also had higher root colonization by mycorrhizal fungi. Endophyte-symbiotic P. leptocoma seeds germinated with greater frequency in P. leptocoma niches relative to P. reflexa niches, whereas neither endophyte-free (experimentally removed) P. leptocoma seeds nor P. reflexa seeds showed differential germination between the two niche types. Thus, endophyte presence constrained the germination and early survival of host plants to microsites occupied by P. leptocoma. However, endophyte-symbiotic P. leptocoma ultimately showed greater growth than endophyte-free plants across all microsites, indicating a net benefit of the symbiosis at this life history stage. Differential effects of endophyte symbiosis on different host life history stages may thus contribute to niche partitioning between the two congeneric plant species. Our study therefore identifies a symbiotic relationship as a potential mechanism facilitating the coexistence of two species, suggesting that symbiont effects on host niche may

  11. Host cells and methods for producing isoprenyl alkanoates

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Taek Soon; Fortman, Jeffrey L.; Keasling, Jay D.

    2015-12-01

    The invention provides for a method of producing an isoprenyl alkanoate in a genetically modified host cell. In one embodiment, the method comprises culturing a genetically modified host cell which expresses an enzyme capable of catalyzing the esterification of an isoprenol and a straight-chain fatty acid, such as an alcohol acetyltransferase (AAT), wax ester synthase/diacylglycerol acyltransferase (WS/DGAT) or lipase, under a suitable condition so that the isoprenyl alkanoate is produced.

  12. Modulation of host-cell MAPkinase signaling during fungal infection

    OpenAIRE

    Nir Osherov

    2015-01-01

    Fungal infections contribute substantially to human suffering and mortality. The interaction between fungal pathogens and their host involves the invasion and penetration of the surface epithelium, activation of cells of the innate immune system and the generation of an effective response to block infection. Numerous host-cell signaling pathways are activated during fungal infection. This review will focus on the main fungal pathogens Aspergillus fumigatus, Candida albicans and Cryptococcus n...

  13. Intracellular Theileria annulata promote invasive cell motility through kinase regulation of the host actin cytoskeleton.

    Directory of Open Access Journals (Sweden)

    Min Ma

    2014-03-01

    Full Text Available The intracellular, protozoan Theileria species parasites are the only eukaryotes known to transform another eukaryotic cell. One consequence of this parasite-dependent transformation is the acquisition of motile and invasive properties of parasitized cells in vitro and their metastatic dissemination in the animal, which causes East Coast Fever (T. parva or Tropical Theileriosis (T. annulata. These motile and invasive properties of infected host cells are enabled by parasite-dependent, poorly understood F-actin dynamics that control host cell membrane protrusions. Herein, we dissected functional and structural alterations that cause acquired motility and invasiveness of T. annulata-infected cells, to understand the molecular basis driving cell dissemination in Tropical Theileriosis. We found that chronic induction of TNFα by the parasite contributes to motility and invasiveness of parasitized host cells. We show that TNFα does so by specifically targeting expression and function of the host proto-oncogenic ser/thr kinase MAP4K4. Blocking either TNFα secretion or MAP4K4 expression dampens the formation of polar, F-actin-rich invasion structures and impairs cell motility in 3D. We identified the F-actin binding ERM family proteins as MAP4K4 downstream effectors in this process because TNFα-induced ERM activation and cell invasiveness are sensitive to MAP4K4 depletion. MAP4K4 expression in infected cells is induced by TNFα-JNK signalling and maintained by the inhibition of translational repression, whereby both effects are parasite dependent. Thus, parasite-induced TNFα promotes invasive motility of infected cells through the activation of MAP4K4, an evolutionary conserved kinase that controls cytoskeleton dynamics and cell motility. Hence, MAP4K4 couples inflammatory signaling to morphodynamic processes and cell motility, a process exploited by the intracellular Theileria parasite to increase its host cell's dissemination capabilities.

  14. Cryptosporidia: epicellular parasites embraced by the host cell membrane.

    Science.gov (United States)

    Valigurová, Andrea; Jirků, Miloslav; Koudela, Bretislav; Gelnar, Milan; Modrý, David; Slapeta, Jan

    2008-07-01

    The ultrastructure of two gastric cryptosporidia, Cryptosporidium muris from experimentally infected rodents (Mastomys natalensis) and Cryptosporidium sp. 'toad' from naturally infected toads (Duttaphrynus melanostictus), was studied using electron microscopy. Observations presented herein allowed us to map ultrastructural aspects of the cryptosporidian invasion process and the origin of a parasitophorous sac. Invading parasites attach to the host cell, followed by gradual envelopment, with the host's cell membrane folds, eventually forming the parasitophorous sac. Cryptosporidian developmental stages remain epicellular during the entire life cycle. The parasite development is illustrated in detail using high resolution field emission scanning electron microscopy. This provides a new insight into the ultrastructural detail of host-parasite interactions and species-specific differences manifested in frequency of detachment of the parasitophorous sac, radial folds of the parasitophorous sac and stem-formation of the parasitised host cell. PMID:18158154

  15. Herpes simplex virus virion host shutoff (vhs) activity alters periocular disease in mice.

    Science.gov (United States)

    Smith, T J; Ackland-Berglund, C E; Leib, D A

    2000-04-01

    During lytic infection, the virion host shutoff (vhs) protein of herpes simplex virus (HSV) mediates the rapid degradation of RNA and shutoff of host protein synthesis. In mice, HSV type 1 (HSV-1) mutants lacking vhs activity are profoundly attenuated. HSV-2 has significantly higher vhs activity than HSV-1, eliciting a faster and more complete shutoff. To examine further the role of vhs activity in pathogenesis, we generated an intertypic recombinant virus (KOSV2) in which the vhs open reading frame of HSV-1 strain KOS was replaced with that of HSV-2 strain 333. KOSV2 and a marker-rescued virus, KOSV2R, were characterized in cell culture and tested in an in vivo mouse eye model of latency and pathogenesis. The RNA degradation kinetics of KOSV2 was identical to that of HSV-2 333, and both showed vhs activity significantly higher than that of KOS. This demonstrated that the fast vhs-mediated degradation phenotype of 333 had been conferred upon KOS. The growth of KOSV2 was comparable to that of KOS, 333, and KOSV2R in cell culture, murine corneas, and trigeminal ganglia and had a reactivation frequency similar to those of KOS and KOSV2R from explanted latently infected trigeminal ganglia. There was, however, significantly reduced blepharitis and viral replication within the periocular skin of KOSV2-infected mice compared to mice infected with either KOS or KOSV2R. Taken together, these data demonstrate that heightened vhs activity, in the context of HSV-1 infection, leads to increased viral clearance from the skin of mice and that the replication of virus in the skin is a determining factor for blepharitis. These data also suggest a role for vhs in modulating host responses to HSV infection.

  16. Brucella abortus Choloylglycine Hydrolase Affects Cell Envelope Composition and Host Cell Internalization

    Science.gov (United States)

    Marchesini, María Inés; Connolly, Joseph; Delpino, María Victoria; Baldi, Pablo C.; Mujer, Cesar V.; DelVecchio, Vito G.; Comerci, Diego J.

    2011-01-01

    Choloylglycine hydrolase (CGH, E.C. 3.5.1.24) is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh) and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization. PMID:22174816

  17. Hyperglycaemia Alters Thymic Epithelial Cell Function

    Directory of Open Access Journals (Sweden)

    Vera Alexandrovna Abramova

    2013-07-01

    Full Text Available Insulin-dependent diabetes mellitus (IDDM is considered to be a consequence of unchecked auto-immune processes. Alterations in immune system responses are thought to be the cause of the disease, but the possibility that altered metabolite levels (glucose can establish the disease by specifically acting on and altering thymus stroma functions has not been investigated. Therefore, the direct effect of hyperglycaemia (HG on central tolerance mechanisms as a causative agent needs to be investigated.

  18. Host epithelial geometry regulates breast cancer cell invasiveness

    Science.gov (United States)

    Boghaert, Eline; Gleghorn, Jason P.; Lee, KangAe; Gjorevski, Nikolce; Radisky, Derek C.; Nelson, Celeste M.

    2012-01-01

    Breast tumor development is regulated in part by cues from the local microenvironment, including interactions with neighboring nontumor cells as well as the ECM. Studies using homogeneous populations of breast cancer cell lines cultured in 3D ECM have shown that increased ECM stiffness stimulates tumor cell invasion. However, at early stages of breast cancer development, malignant cells are surrounded by normal epithelial cells, which have been shown to exert a tumor-suppressive effect on cocultured cancer cells. Here we explored how the biophysical characteristics of the host microenvironment affect the proliferative and invasive tumor phenotype of the earliest stages of tumor development, by using a 3D microfabrication-based approach to engineer ducts composed of normal mammary epithelial cells that contained a single tumor cell. We found that the phenotype of the tumor cell was dictated by its position in the duct: proliferation and invasion were enhanced at the ends and blocked when the tumor cell was located elsewhere within the tissue. Regions of invasion correlated with high endogenous mechanical stress, as shown by finite element modeling and bead displacement experiments, and modulating the contractility of the host epithelium controlled the subsequent invasion of tumor cells. Combining microcomputed tomographic analysis with finite element modeling suggested that predicted regions of high mechanical stress correspond to regions of tumor formation in vivo. This work suggests that the mechanical tone of nontumorigenic host epithelium directs the phenotype of tumor cells and provides additional insight into the instructive role of the mechanical tumor microenvironment. PMID:23150585

  19. Control of Host Cell Phosphorylation by Legionella Pneumophila

    OpenAIRE

    Haenssler, Eva; Isberg, Ralph R.

    2011-01-01

    Phosphorylation is one of the most frequent modifications in intracellular signaling and is implicated in many processes ranging from transcriptional control to signal transduction in innate immunity. Many pathogens modulate host cell phosphorylation pathways to promote growth and establish an infectious disease. The intracellular pathogen Legionella pneumophila targets and exploits the host phosphorylation system throughout the infection cycle as part of its strategy to establish an environm...

  20. Transcriptome and microRNome of Theileria annulata Host Cells

    KAUST Repository

    Rchiad, Zineb

    2016-06-01

    Tropical Theileriosis is a parasitic disease of calves with a profound economic impact caused by Theileria annulata, an apicomplexan parasite of the genus Theileria. Transmitted by Hyalomma ticks, T. annulata infects and transforms bovine lymphocytes and macrophages into a cancer-like phenotype characterized by all six hallmarks of cancer. In the current study we investigate the transcriptional landscape of T. annulata-infected lymphocytes to define genes and miRNAs regulated by host cell transformation using next generation sequencing. We also define genes and miRNAs differentially expressed as a result of the attenuation of a T.annulata-infected macrophage cell line used as a vaccine. By comparing the transcriptional landscape of one attenuated and two transformed cell lines we identify four genes that we propose as key factors in transformation and virulence of the T. annulata host cells. We also identify miR- 126-5p as a key regulator of infected cells proliferation, adhesion, survival and invasiveness. In addition to the host cell trascriptome we studied T. annulata transcriptome and identified the role of ROS and TGF-β2 in controlling parasite gene expression. Moreover, we have used the deep parasite ssRNA-seq data to refine the available T. annulata annotation. Taken together, this study provides the full list of host cell’s genes and miRNAs transcriptionally perturbed after infection with T. annulata and after attenuation and describes genes and miRNAs never identified before as players in this type of host cell transformation. Moreover, this study provides the first database for the transcriptome of T. annulata and its host cells using next generation sequencing.

  1. Aquatic viruses induce host cell death pathways and its application.

    Science.gov (United States)

    Reshi, Latif; Wu, Jen-Leih; Wang, Hao-Ven; Hong, Jiann-Ruey

    2016-01-01

    Virus infections of mammalian and animal cells consist of a series of events. As intracellular parasites, viruses rely on the use of host cellular machinery. Through the use of cell culture and molecular approaches over the past decade, our knowledge of the biology of aquatic viruses has grown exponentially. The increase in aquaculture operations worldwide has provided new approaches for the transmission of aquatic viruses that include RNA and DNA viruses. Therefore, the struggle between the virus and the host for control of the cell's death machinery is crucial for survival. Viruses are obligatory intracellular parasites and, as such, must modulate apoptotic pathways to control the lifespan of their host to complete their replication cycle. This paper updates the discussion on the detailed mechanisms of action that various aquatic viruses use to induce cell death pathways in the host, such as Bad-mediated, mitochondria-mediated, ROS-mediated and Fas-mediated cell death circuits. Understanding how viruses exploit the apoptotic pathways of their hosts may provide great opportunities for the development of future potential therapeutic strategies and pathogenic insights into different aquatic viral diseases.

  2. Malaria: targeting parasite and host cell kinomes.

    Science.gov (United States)

    Doerig, Christian; Abdi, Abdirahman; Bland, Nicholas; Eschenlauer, Sylvain; Dorin-Semblat, Dominique; Fennell, Clare; Halbert, Jean; Holland, Zoe; Nivez, Marie-Paule; Semblat, Jean-Philippe; Sicard, Audrey; Reininger, Luc

    2010-03-01

    Malaria still remains one of the deadliest infectious diseases, and has a tremendous morbidity and mortality impact in the developing world. The propensity of the parasites to develop drug resistance, and the relative reluctance of the pharmaceutical industry to invest massively in the developments of drugs that would offer only limited marketing prospects, are major issues in antimalarial drug discovery. Protein kinases (PKs) have become a major family of targets for drug discovery research in a number of disease contexts, which has generated considerable resources such as kinase-directed libraries and high throughput kinase inhibition assays. The phylogenetic distance between malaria parasites and their human host translates into important divergences in their respective kinomes, and most Plasmodium kinases display atypical properties (as compared to mammalian PKs) that can be exploited towards selective inhibition. Here, we discuss the taxon-specific kinases possessed by malaria parasites, and give an overview of target PKs that have been validated by reverse genetics, either in the human malaria parasite Plasmodium falciparum or in the rodent model Plasmodium berghei. We also briefly allude to the possibility of attacking Plasmodium through the inhibition of human PKs that are required for survival of this obligatory intracellular parasite, and which are targets for other human diseases. PMID:19840874

  3. Adaptive immunity alters distinct host feeding pathways during nematode induced inflammation, a novel mechanism in parasite expulsion.

    Directory of Open Access Journals (Sweden)

    John J Worthington

    2013-01-01

    Full Text Available Gastrointestinal infection is often associated with hypophagia and weight loss; however, the precise mechanisms governing these responses remain poorly defined. Furthermore, the possibility that alterations in feeding during infection may be beneficial to the host requires further study. We used the nematode Trichinella spiralis, which transiently inhabits the small intestine before migrating to skeletal muscle, as a biphasic model of infection to determine the cellular and molecular pathways controlling feeding during enteric and peripheral inflammation. Through the infection of genetically modified mice lacking cholecystokinin, Tumor necrosis factor α receptors and T and B-cells, we observed a biphasic hypophagic response to infection resulting from two separate immune-driven mechanisms. The enteroendocrine I-cell derived hormone cholecystokinin is an essential mediator of initial hypophagia and is induced by CD4+ T-cells during enteritis. In contrast, the second hypophagic response is extra-intestinal and due to the anorectic effects of TNFα during peripheral infection of the muscle. Moreover, via maintaining naive levels of the adipose secreted hormone leptin throughout infection we demonstrate a novel feedback loop in the immunoendocrine axis. Immune driven I-cell hyperplasia and resultant weight loss leads to a reduction in the inflammatory adipokine leptin, which in turn heightens protective immunity during infection. These results characterize specific immune mediated mechanisms which reduce feeding during intestinal or peripheral inflammation. Importantly, the molecular mediators of each phase are entirely separate. The data also introduce the first evidence that I-cell hyperplasia is an adaptively driven immune response that directly impinges on the outcome to infection.

  4. Cytokine-dependent and–independent gene expression changes and cell cycle block revealed in Trypanosoma cruzi-infected host cells by comparative mRNA profiling

    Directory of Open Access Journals (Sweden)

    Burleigh Barbara A

    2009-05-01

    Full Text Available Abstract Background The requirements for growth and survival of the intracellular pathogen Trypanosoma cruzi within mammalian host cells are poorly understood. Transcriptional profiling of the host cell response to infection serves as a rapid read-out for perturbation of host physiology that, in part, reflects adaptation to the infective process. Using Affymetrix oligonucleotide array analysis we identified common and disparate host cell responses triggered by T. cruzi infection of phenotypically diverse human cell types. Results We report significant changes in transcript abundance in T. cruzi-infected fibroblasts, endothelial cells and smooth muscle cells (2852, 2155 and 531 genes respectively; fold-change ≥ 2, p-value T. cruzi-infected fibroblasts and endothelial cells transwell plates were used to distinguish cytokine-dependent and -independent gene expression profiles. This approach revealed the induction of metabolic and signaling pathways involved in cell proliferation, amino acid catabolism and response to wounding as common themes in T. cruzi-infected cells. In addition, the downregulation of genes involved in mitotic cell cycle and cell division predicted that T. cruzi infection may impede host cell cycle progression. The observation of impaired cytokinesis in T. cruzi-infected cells, following nuclear replication, confirmed this prediction. Conclusion Metabolic pathways and cellular processes were identified as significantly altered at the transcriptional level in response to T. cruzi infection in a cytokine-independent manner. Several of these alterations are supported by previous studies of T. cruzi metabolic requirements or effects on the host. However, our methods also revealed a T. cruzi-dependent block in the host cell cycle, at the level of cytokinesis, previously unrecognized for this pathogen-host cell interaction.

  5. Bicarbonate correction of ketoacidosis alters host-pathogen interactions and alleviates mucormycosis.

    Science.gov (United States)

    Gebremariam, Teclegiorgis; Lin, Lin; Liu, Mingfu; Kontoyiannis, Dimitrios P; French, Samuel; Edwards, John E; Filler, Scott G; Ibrahim, Ashraf S

    2016-06-01

    Patients with diabetic ketoacidosis (DKA) are uniquely predisposed to mucormycosis, an angioinvasive fungal infection with high mortality. Previously, we demonstrated that Rhizopus invades the endothelium via binding of fungal CotH proteins to the host receptor GRP78. Here, we report that surface expression of GRP78 is increased in endothelial cells exposed to physiological concentrations of β-hydroxy butyrate (BHB), glucose, and iron that are similar to those found in DKA patients. Additionally, expression of R. oryzae CotH was increased within hours of incubation with DKA-associated concentrations of BHB, glucose, and iron, augmenting the ability of R. oryzae to invade and subsequently damage endothelial cells in vitro. BHB exposure also increased fungal growth and attenuated R. oryzae neutrophil-mediated damage. Further, mice given BHB developed clinical acidosis and became extremely susceptible to mucormycosis, but not aspergillosis, while sodium bicarbonate reversed this susceptibility. BHB-related acidosis exerted a direct effect on both GRP78 and CotH expression, an effect not seen with lactic acidosis. However, BHB also indirectly compromised the ability of transferrin to chelate iron, as iron chelation combined with sodium bicarbonate completely protected endothelial cells from Rhizopus-mediated invasion and damage. Our results dissect the pathogenesis of mucormycosis during ketoacidosis and reinforce the importance of careful metabolic control of the acidosis to prevent and manage this infection. PMID:27159390

  6. Early Bunyavirus-Host Cell Interactions

    Directory of Open Access Journals (Sweden)

    Amelina Albornoz

    2016-05-01

    Full Text Available The Bunyaviridae is the largest family of RNA viruses, with over 350 members worldwide. Several of these viruses cause severe diseases in livestock and humans. With an increasing number and frequency of outbreaks, bunyaviruses represent a growing threat to public health and agricultural productivity globally. Yet, the receptors, cellular factors and endocytic pathways used by these emerging pathogens to infect cells remain largely uncharacterized. The focus of this review is on the early steps of bunyavirus infection, from virus binding to penetration from endosomes. We address current knowledge and advances for members from each genus in the Bunyaviridae family regarding virus receptors, uptake, intracellular trafficking and fusion.

  7. HumanViCe: Host ceRNA network in virus infected cells in human

    Directory of Open Access Journals (Sweden)

    Suman eGhosal

    2014-07-01

    Full Text Available Host-virus interaction via host cellular components has been an important field of research in recent times. RNA interference mediated by short interfering RNAs and microRNAs (miRNA, is a widespread anti-viral defence strategy. Importantly, viruses also encode their own miRNAs. In recent times miRNAs were identified as key players in host-virus interaction. Furthermore, viruses were shown to exploit the host miRNA networks to suite their own need. The complex cross-talk between host and viral miRNAs and their cellular and viral targets forms the environment for viral pathogenesis. Apart from protein-coding mRNAs, non-coding RNAs may also be targeted by host or viral miRNAs in virus infected cells, and viruses can exploit the host miRNA mediated gene regulatory network via the competing endogenous RNA effect. A recent report showed that viral U-rich non-coding RNAs called HSUR, expressed in primate virus herpesvirus saimiri (HVS infected T cells, were able to bind to three host miRNAs, causing significant alteration in cellular level for one of the miRNAs. We have predicted protein coding and non protein-coding targets for viral and human miRNAs in virus infected cells. We identified viral miRNA targets within host non-coding RNA loci from AGO interacting regions in three different virus infected cells. Gene ontology (GO and pathway enrichment analysis of the genes comprising the ceRNA networks in the virus infected cells revealed enrichment of key cellular signalling pathways related to cell fate decisions and gene transcription, like Notch and Wnt signalling pathways, as well as pathways related to viral entry, replication and virulence. We identified a vast number of non-coding transcripts playing as potential ceRNAs to the immune response associated genes; e.g. APOBEC family genes, in some virus infected cells. All these information are compiled in HumanViCe, a comprehensive database that provides the potential ceRNA networks in virus

  8. Lipid exchange between Borrelia burgdorferi and host cells.

    Directory of Open Access Journals (Sweden)

    Jameson T Crowley

    2013-01-01

    Full Text Available Borrelia burgdorferi, the agent of Lyme disease, has cholesterol and cholesterol-glycolipids that are essential for bacterial fitness, are antigenic, and could be important in mediating interactions with cells of the eukaryotic host. We show that the spirochetes can acquire cholesterol from plasma membranes of epithelial cells. In addition, through fluorescent and confocal microscopy combined with biochemical approaches, we demonstrated that B. burgdorferi labeled with the fluorescent cholesterol analog BODIPY-cholesterol or (3H-labeled cholesterol transfer both cholesterol and cholesterol-glycolipids to HeLa cells. The transfer occurs through two different mechanisms, by direct contact between the bacteria and eukaryotic cell and/or through release of outer membrane vesicles. Thus, two-way lipid exchange between spirochetes and host cells can occur. This lipid exchange could be an important process that contributes to the pathogenesis of Lyme disease.

  9. Knockdown of Five Genes Encoding Uncharacterized Proteins Inhibits Entamoeba histolytica Phagocytosis of Dead Host Cells.

    Science.gov (United States)

    Sateriale, Adam; Miller, Peter; Huston, Christopher D

    2016-04-01

    Entamoeba histolytica is the protozoan parasite that causes invasive amebiasis, which is endemic to many developing countries and characterized by dysentery and liver abscesses. The virulence of E. histolytica correlates with the degree of host cell engulfment, or phagocytosis, and E. histolytica phagocytosis alters amebic gene expression in a feed-forward manner that results in an increased phagocytic ability. Here, we used a streamlined RNA interference screen to silence the expression of 15 genes whose expression was upregulated in phagocytic E. histolytica trophozoites to determine whether these genes actually function in the phagocytic process. When five of these genes were silenced, amebic strains with significant decreases in the ability to phagocytose apoptotic host cells were produced. Phagocytosis of live host cells, however, was largely unchanged, and the defects were surprisingly specific for phagocytosis. Two of the five encoded proteins, which we named E. histolytica ILWEQ (EhILWEQ) and E. histolytica BAR (EhBAR), were chosen for localization via SNAP tag labeling and localized to the site of partially formed phagosomes. Therefore, both EhILWEQ and EhBAR appear to contribute to E. histolytica virulence through their function in phagocytosis, and the large proportion (5/15 [33%]) of gene-silenced strains with a reduced ability to phagocytose host cells validates the previously published microarray data set demonstrating feed-forward control of E. histolytica phagocytosis. Finally, although only limited conclusions can be drawn from studies using the virulence-deficient G3 Entamoeba strain, the relative specificity of the defects induced for phagocytosis of apoptotic cells but not healthy cells suggests that cell killing may play a rate-limiting role in the process of Entamoeba histolytica host cell engulfment. PMID:26810036

  10. Knockdown of Five Genes Encoding Uncharacterized Proteins Inhibits Entamoeba histolytica Phagocytosis of Dead Host Cells.

    Science.gov (United States)

    Sateriale, Adam; Miller, Peter; Huston, Christopher D

    2016-04-01

    Entamoeba histolytica is the protozoan parasite that causes invasive amebiasis, which is endemic to many developing countries and characterized by dysentery and liver abscesses. The virulence of E. histolytica correlates with the degree of host cell engulfment, or phagocytosis, and E. histolytica phagocytosis alters amebic gene expression in a feed-forward manner that results in an increased phagocytic ability. Here, we used a streamlined RNA interference screen to silence the expression of 15 genes whose expression was upregulated in phagocytic E. histolytica trophozoites to determine whether these genes actually function in the phagocytic process. When five of these genes were silenced, amebic strains with significant decreases in the ability to phagocytose apoptotic host cells were produced. Phagocytosis of live host cells, however, was largely unchanged, and the defects were surprisingly specific for phagocytosis. Two of the five encoded proteins, which we named E. histolytica ILWEQ (EhILWEQ) and E. histolytica BAR (EhBAR), were chosen for localization via SNAP tag labeling and localized to the site of partially formed phagosomes. Therefore, both EhILWEQ and EhBAR appear to contribute to E. histolytica virulence through their function in phagocytosis, and the large proportion (5/15 [33%]) of gene-silenced strains with a reduced ability to phagocytose host cells validates the previously published microarray data set demonstrating feed-forward control of E. histolytica phagocytosis. Finally, although only limited conclusions can be drawn from studies using the virulence-deficient G3 Entamoeba strain, the relative specificity of the defects induced for phagocytosis of apoptotic cells but not healthy cells suggests that cell killing may play a rate-limiting role in the process of Entamoeba histolytica host cell engulfment.

  11. Colonic microbiota alters host susceptibility to infectious colitis by modulating inflammation, redox status, and ion transporter gene expression.

    Science.gov (United States)

    Ghosh, S; Dai, C; Brown, K; Rajendiran, E; Makarenko, S; Baker, J; Ma, C; Halder, S; Montero, M; Ionescu, V A; Klegeris, A; Vallance, B A; Gibson, D L

    2011-07-01

    Individuals vary in their resistance to enteric infections. The role of the intestinal microbiota in altering susceptibility to enteric infection is relatively unknown. Previous studies have identified that C3H/HeOuJ mice suffer 100% mortality during Citrobacter rodentium-induced colitis, whereas C57BL/6 mice recover from infection. The basis for their differences in susceptibility is unclear and has been mainly attributed to differences in host genetics. This study investigated the role of the intestinal microbiota in altering susceptibility to C. rodentium-induced colitis. When the feces of C57BL/6 mice were gavaged into antibiotic treated C3H/HeOuJ mice, the C57BL/6 microflora led to a complete reversal in mortality patterns where 100% of the C3H/HeOuJ mice survived infection. This protection corresponded with reduced colonic pathology and less systemic pathogen load and was associated with increased inflammatory and redox responses with reduced epithelial cell death. C3H/HeOuJ mice are normally susceptible to infection-induced dehydration due to defective expression of colonic ion transporters such as Dra, CA IV, and CA I; expression of these genes was normalized when C3H/HeOuJ mice were colonized with the C57BL/6 microflora. Together, these data reveal that the colonic microbiota play a critical role in protecting against intestinal infection by inducing proinflammatory and prooxidant responses that control pathogen load as well as ion transporter gene expression previously shown to prevent fatal dehydration. Protection of mice from lethal colitis was associated with higher levels of bacteria from Bacteroidetes. This study reveals that the microbiota is sufficient to overcome inherent genetic susceptibility patterns in C3H/HeOuJ mice that cause mortality during C. rodentium infection.

  12. Plasmodium species: master renovators of their host cells.

    Science.gov (United States)

    de Koning-Ward, Tania F; Dixon, Matthew W A; Tilley, Leann; Gilson, Paul R

    2016-08-01

    Plasmodium parasites, the causative agents of malaria, have developed elaborate strategies that they use to survive and thrive within different intracellular environments. During the blood stage of infection, the parasite is a master renovator of its erythrocyte host cell, and the changes in cell morphology and function that are induced by the parasite promote survival and contribute to the pathogenesis of severe malaria. In this Review, we discuss how Plasmodium parasites use the protein trafficking motif Plasmodium export element (PEXEL), protease-mediated polypeptide processing, a novel translocon termed the Plasmodium translocon of exported proteins (PTEX) and exomembranous structures to export hundreds of proteins to discrete subcellular locations in the host erythrocytes, which enables the parasite to gain access to vital nutrients and to evade the immune defence mechanisms of the host.

  13. Plasmodium species: master renovators of their host cells.

    Science.gov (United States)

    de Koning-Ward, Tania F; Dixon, Matthew W A; Tilley, Leann; Gilson, Paul R

    2016-08-01

    Plasmodium parasites, the causative agents of malaria, have developed elaborate strategies that they use to survive and thrive within different intracellular environments. During the blood stage of infection, the parasite is a master renovator of its erythrocyte host cell, and the changes in cell morphology and function that are induced by the parasite promote survival and contribute to the pathogenesis of severe malaria. In this Review, we discuss how Plasmodium parasites use the protein trafficking motif Plasmodium export element (PEXEL), protease-mediated polypeptide processing, a novel translocon termed the Plasmodium translocon of exported proteins (PTEX) and exomembranous structures to export hundreds of proteins to discrete subcellular locations in the host erythrocytes, which enables the parasite to gain access to vital nutrients and to evade the immune defence mechanisms of the host. PMID:27374802

  14. Host cell P-glycoprotein is essential for cholesterol uptake and replication of Toxoplasma gondii.

    Science.gov (United States)

    Bottova, Iveta; Hehl, Adrian B; Stefanić, Sasa; Fabriàs, Gemma; Casas, Josefina; Schraner, Elisabeth; Pieters, Jean; Sonda, Sabrina

    2009-06-26

    P-glycoprotein (P-gp) is a membrane-bound efflux pump that actively exports a wide range of compounds from the cell and is associated with the phenomenon of multidrug resistance. However, the role of P-gp in normal physiological processes remains elusive. Using P-gp-deficient fibroblasts, we showed that P-gp was critical for the replication of the intracellular parasite Toxoplasma gondii but was not involved in invasion of host cells by the parasite. Importantly, we found that the protein participated in the transport of host-derived cholesterol to the intracellular parasite. T. gondii replication in P-gp-deficient host cells not only resulted in reduced cholesterol content in the parasite but also altered its sphingolipid metabolism. In addition, we found that different levels of P-gp expression modified the cholesterol metabolism in uninfected fibroblasts. Collectively our findings reveal a key and previously undocumented role of P-gp in host-parasite interaction and suggest a physiological role for P-gp in cholesterol trafficking in mammalian cells. PMID:19389707

  15. Host Cell P-glycoprotein Is Essential for Cholesterol Uptake and Replication of Toxoplasma gondii*

    Science.gov (United States)

    Bottova, Iveta; Hehl, Adrian B.; Štefanić, Saša; Fabriàs, Gemma; Casas, Josefina; Schraner, Elisabeth; Pieters, Jean; Sonda, Sabrina

    2009-01-01

    P-glycoprotein (P-gp) is a membrane-bound efflux pump that actively exports a wide range of compounds from the cell and is associated with the phenomenon of multidrug resistance. However, the role of P-gp in normal physiological processes remains elusive. Using P-gp-deficient fibroblasts, we showed that P-gp was critical for the replication of the intracellular parasite Toxoplasma gondii but was not involved in invasion of host cells by the parasite. Importantly, we found that the protein participated in the transport of host-derived cholesterol to the intracellular parasite. T. gondii replication in P-gp-deficient host cells not only resulted in reduced cholesterol content in the parasite but also altered its sphingolipid metabolism. In addition, we found that different levels of P-gp expression modified the cholesterol metabolism in uninfected fibroblasts. Collectively our findings reveal a key and previously undocumented role of P-gp in host-parasite interaction and suggest a physiological role for P-gp in cholesterol trafficking in mammalian cells. PMID:19389707

  16. Clustered Intracellular Salmonella enterica Serovar Typhimurium Blocks Host Cell Cytokinesis.

    Science.gov (United States)

    Santos, António J M; Durkin, Charlotte H; Helaine, Sophie; Boucrot, Emmanuel; Holden, David W

    2016-07-01

    Several bacterial pathogens and viruses interfere with the cell cycle of their host cells to enhance virulence. This is especially apparent in bacteria that colonize the gut epithelium, where inhibition of the cell cycle of infected cells enhances the intestinal colonization. We found that intracellular Salmonella enterica serovar Typhimurium induced the binucleation of a large proportion of epithelial cells by 14 h postinvasion and that the effect was dependent on an intact Salmonella pathogenicity island 2 (SPI-2) type 3 secretion system. The SPI-2 effectors SseF and SseG were required to induce binucleation. SseF and SseG are known to maintain microcolonies of Salmonella-containing vacuoles close to the microtubule organizing center of infected epithelial cells. During host cell division, these clustered microcolonies prevented the correct localization of members of the chromosomal passenger complex and mitotic kinesin-like protein 1 and consequently prevented cytokinesis. Tetraploidy, arising from a cytokinesis defect, is known to have a deleterious effect on subsequent cell divisions, resulting in either chromosomal instabilities or cell cycle arrest. In infected mice, proliferation of small intestinal epithelial cells was compromised in an SseF/SseG-dependent manner, suggesting that cytokinesis failure caused by S Typhimurium delays epithelial cell turnover in the intestine.

  17. Seasonal alterations in host range and fidelity in the polyphagous mirid bug, Apolygus lucorum (Heteroptera: Miridae.

    Directory of Open Access Journals (Sweden)

    Hongsheng Pan

    Full Text Available In herbivorous insects, host plant switching is commonly observed and plays an important role in their annual life cycle. However, much remains to be learned about seasonal host switching of various pestiferous arthropods under natural conditions. From 2006 until 2012, we assessed Apolygus lucorum (Meyer-Dür host plant use in successive spring, summer and winter seasons at one single location (Langfang, China. Data were used to quantify changes in host plant breadth and host fidelity between seasons. Host fidelity of A. lucorum differed between seasons, with 87.9% of spring hosts also used in the summer and 36.1% of summer hosts used in winter. In contrast, as little as 25.6% host plant species were shared between winter and spring. Annual herbaceous plants are most often used for overwintering, while perennial woody plants are relatively important for initial population build-up in the spring. Our study contributes to an improved understanding of evolutionary interactions between A. lucorum and its host plants and lays the groundwork for the design of population management strategies for this important pest in myriad crops.

  18. Implanted neural progenitor cells regulate glial reaction to brain injury and establish gap junctions with host glial cells.

    Science.gov (United States)

    Talaverón, Rocío; Matarredona, Esperanza R; de la Cruz, Rosa R; Macías, David; Gálvez, Victoria; Pastor, Angel M

    2014-04-01

    Transplantation of neural stem/progenitor cells (NPCs) in the lesioned brain is able to restore morphological and physiological alterations induced by different injuries. The local microenvironment created at the site of grafting and the communication between grafted and host cells are crucial in the beneficial effects attributed to the NPC implants. We have previously described that NPC transplantation in an animal model of central axotomy restores firing properties and synaptic coverage of lesioned neurons and modulates their trophic factor content. In this study, we aim to explore anatomical relationships between implanted NPCs and host glia that might account for the implant-induced neuroprotective effects. Postnatal rat subventricular zone NPCs were isolated and grafted in adult rats after transection of the medial longitudinal fascicle. Brains were removed and analyzed eight weeks later. Immunohistochemistry for different glial markers revealed that NPC-grafted animals displayed significantly greater microglial activation than animals that received only vehicle injections. Implanted NPCs were located in close apposition to activated microglia and reactive astrocytes. The gap junction protein connexin43 was present in NPCs and glial cells at the lesion site and was often found interposed within adjacent implanted and glial cells. Gap junctions were identified between implanted NPCs and host astrocytes and less frequently between NPCs and microglia. Our results show that implanted NPCs modulate the glial reaction to lesion and establish the possibility of communication through gap junctions between grafted and host glial cells which might be involved in the restorative effects of NPC implants.

  19. Physiological alterations in UV-irradiated cells: liquid holding recovery

    International Nuclear Information System (INIS)

    The biochemical and physiological alterations that occur in ultraviolet irradiated cells, during liquid holding have been studied. Incubation in buffer acts not to interfer directly with the mechanic repairs but by promoting metabolic alterations that would block some irreversible and lethal physiological responses. (L.M.J.)

  20. Necroptosis: The Trojan horse in cell autonomous antiviral host defense.

    Science.gov (United States)

    Mocarski, Edward S; Guo, Hongyan; Kaiser, William J

    2015-05-01

    Herpesviruses suppress cell death to assure sustained infection in their natural hosts. Murine cytomegalovirus (MCMV) encodes suppressors of apoptosis as well as M45-encoded viral inhibitor of RIP activation (vIRA) to block RIP homotypic interaction motif (RHIM)-signaling and recruitment of RIP3 (also called RIPK3), to prevent necroptosis. MCMV and human cytomegalovirus encode a viral inhibitor of caspase (Casp)8 activation to block apoptosis, an activity that unleashes necroptosis. Herpes simplex virus (HSV)1 and HSV2 incorporate both RHIM and Casp8 suppression strategies within UL39-encoded ICP6 and ICP10, respectively, which are herpesvirus-conserved homologs of MCMV M45. Both HSV proteins sensitize human cells to necroptosis by blocking Casp8 activity while preventing RHIM-dependent RIP3 activation and death. In mouse cells, HSV1 ICP6 interacts with RIP3 and, surprisingly, drives necroptosis. Thus, herpesviruses have illuminated the contribution of necoptosis to host defense in the natural host as well as its potential to restrict cross-species infections in nonnatural hosts. PMID:25819165

  1. Raman spectroscopic study of a genetically altered kidney cell

    Science.gov (United States)

    Joshi, Joel; Garcia, Francisco; Centeno, Silvia P.; Joshi, N. V.

    2008-02-01

    A Raman spectroscopic investigation of a genetically altered Human Embryonic Kidney Cell (HEK293) along with a pathologically normal cell has been carried out by a conventional method. The genetic alteration was carried out with a standard protocol by using a Green Fluorescence Protein (GFP). Raman spectra show that there are dramatic differences between the spectrum obtained from a genetically altered cell and that obtained from a pathologically normal cell. The former shows three broad bands; meanwhile the latter shows several sharp peaks corresponding to the ring vibrational modes of Phen, GFP and DNA. The present analysis provides an indication that the force field near Phen located at 64, 65 and 66 was altered during the genetic transformation. The Raman spectrum could be a direct experimental evidence for substantial modifications triggered due to the expression of specific genes.

  2. Climate Change May Alter Breeding Ground Distributions of Eastern Migratory Monarchs (Danaus plexippus) via Range Expansion of Asclepias Host Plants

    OpenAIRE

    Nathan P Lemoine

    2015-01-01

    Climate change can profoundly alter species’ distributions due to changes in temperature, precipitation, or seasonality. Migratory monarch butterflies (Danaus plexippus) may be particularly susceptible to climate-driven changes in host plant abundance or reduced overwintering habitat. For example, climate change may significantly reduce the availability of overwintering habitat by restricting the amount of area with suitable microclimate conditions. However, potential effects of climate chang...

  3. Recombinant host cells and media for ethanol production

    Science.gov (United States)

    Wood, Brent E; Ingram, Lonnie O; Yomano, Lorraine P; York, Sean W

    2014-02-18

    Disclosed are recombinant host cells suitable for degrading an oligosaccharide that have been optimized for growth and production of high yields of ethanol, and methods of making and using these cells. The invention further provides minimal media comprising urea-like compounds for economical production of ethanol by recombinant microorganisms. Recombinant host cells in accordance with the invention are modified by gene mutation to eliminate genes responsible for the production of unwanted products other than ethanol, thereby increasing the yield of ethanol produced from the oligosaccharides, relative to unmutated parent strains. The new and improved strains of recombinant bacteria are capable of superior ethanol productivity and yield when grown under conditions suitable for fermentation in minimal growth media containing inexpensive reagents. Systems optimized for ethanol production combine a selected optimized minimal medium with a recombinant host cell optimized for use in the selected medium. Preferred systems are suitable for efficient ethanol production by simultaneous saccharification and fermentation (SSF) using lignocellulose as an oligosaccharide source. The invention also provides novel isolated polynucleotide sequences, polypeptide sequences, vectors and antibodies.

  4. Alteration of cell cycle progression by Sindbis virus infection

    Energy Technology Data Exchange (ETDEWEB)

    Yi, Ruirong; Saito, Kengo [Department of Molecular Virology, Graduate School of Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670 (Japan); Isegawa, Naohisa [Laboratory Animal Center, Graduate School of Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670 (Japan); Shirasawa, Hiroshi, E-mail: sirasawa@faculty.chiba-u.jp [Department of Molecular Virology, Graduate School of Medicine, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chiba 260-8670 (Japan)

    2015-07-10

    We examined the impact of Sindbis virus (SINV) infection on cell cycle progression in a cancer cell line, HeLa, and a non-cancerous cell line, Vero. Cell cycle analyses showed that SINV infection is able to alter the cell cycle progression in both HeLa and Vero cells, but differently, especially during the early stage of infection. SINV infection affected the expression of several cell cycle regulators (CDK4, CDK6, cyclin E, p21, cyclin A and cyclin B) in HeLa cells and caused HeLa cells to accumulate in S phase during the early stage of infection. Monitoring SINV replication in HeLa and Vero cells expressing cell cycle indicators revealed that SINV which infected HeLa cells during G{sub 1} phase preferred to proliferate during S/G{sub 2} phase, and the average time interval for viral replication was significantly shorter in both HeLa and Vero cells infected during G{sub 1} phase than in cells infected during S/G{sub 2} phase. - Highlights: • SINV infection was able to alter the cell cycle progression of infected cancer cells. • SINV infection can affect the expression of cell cycle regulators. • SINV infection exhibited a preference for the timing of viral replication among the cell cycle phases.

  5. Infection strategies of intestinal parasite pathogens and host cell responses

    Directory of Open Access Journals (Sweden)

    Bruno Martorell Di Genova

    2016-03-01

    Full Text Available Giardia lamblia, Cryptosporidium spp. and Entamoeba histolytica are important pathogenic intestinal parasites and are amongst the leading cause worldwide of diarrheal illness in humans. Diseases caused by these organisms, Giardiasis, Cryptosporidiosis and Amoebiasis, respectively, are characterized by self-limited diarrhea but can evolve to long-term complications. The cellular and molecular mechanisms underlying the pathogenesis of diarrhea associated with these tree pathogens are being unraveled, with knowledge of both the strategies explored by the parasites to establish infection and the methods evolved by hosts to avoid it. Special attention is being given to molecules participating in parasite-host interaction and in the mechanisms implicated in the diseases pathophysiologic processes. This review focuses on cell mechanisms that are modulated during infection, including gene transcription, cytoskeleton rearrangements, signal transduction pathways and cell death.

  6. Brucella T4SS: the VIP pass inside host cells.

    Science.gov (United States)

    Lacerda, Thais Lourdes Santos; Salcedo, Suzana Pinto; Gorvel, Jean-Pierre

    2013-02-01

    For many Gram-negative bacteria, like Brucella, the type IV secretion system (T4SS) has a critical role in bacterial virulence. In Brucella, the VirB T4SS permits the injection of bacterial effectors inside host cells, leading to subversion of signaling pathways and favoring bacterial growth and pathogenesis. The virB operon promoter is tightly regulated by a combination of transcriptional activators and repressors that are expressed according to the environmental conditions encountered by Brucella. Recent advances have shed light on the Brucella T4SS regulatory mechanisms and also its substrates. Characterization of the targets and functions of these translocated effectors is underway and will help understand the role of the T4SS in the establishment of a replication niche inside host cells.

  7. Infection Strategies of Intestinal Parasite Pathogens and Host Cell Responses.

    Science.gov (United States)

    Di Genova, Bruno M; Tonelli, Renata R

    2016-01-01

    Giardia lamblia, Cryptosporidium sp., and Entamoeba histolytica are important pathogenic intestinal parasites and are amongst the leading causes worldwide of diarrheal illness in humans. Diseases caused by these organisms, giardiasis, cryptosporidiosis, and amoebiasis, respectively, are characterized by self-limited diarrhea but can evolve to long-term complications. The cellular and molecular mechanisms underlying the pathogenesis of diarrhea associated with these three pathogens are being unraveled, with knowledge of both the strategies explored by the parasites to establish infection and the methods evolved by hosts to avoid it. Special attention is being given to molecules participating in parasite-host interaction and in the mechanisms implicated in the diseases' pathophysiologic processes. This review focuses on cell mechanisms that are modulated during infection, including gene transcription, cytoskeleton rearrangements, signal transduction pathways, and cell death. PMID:26973630

  8. Host Cell Autophagy in Immune Response to Zoonotic Infections

    Directory of Open Access Journals (Sweden)

    Panagiotis Skendros

    2012-01-01

    Full Text Available Autophagy is a fundamental homeostatic process in which cytoplasmic targets are sequestered within double-membraned autophagosomes and subsequently delivered to lysosomes for degradation. Accumulating evidence supports the pivotal role of autophagy in host defense against intracellular pathogens implicating both innate and adaptive immunity. Many of these pathogens cause common zoonotic infections worldwide. The induction of the autophagic machinery by innate immune receptors signaling, such as TLRs, NOD1/2, and p62/SQSTM1 in antigen-presenting cells results in inhibition of survival and elimination of invading pathogens. Furthermore, Th1 cytokines induce the autophagic process, whereas autophagy also contributes to antigen processing and MHC class II presentation, linking innate to adaptive immunity. However, several pathogens have developed strategies to avoid autophagy or exploit autophagic machinery to their advantage. This paper focuses on the role of host cell autophagy in the regulation of immune response against intracellular pathogens, emphasizing on selected bacterial and protozoan zoonoses.

  9. Uropathogenic Escherichia coli Epigenetically Manipulate Host Cell Death Pathways.

    Science.gov (United States)

    Zhang, Zhengguo; Wang, Ming; Eisel, Florian; Tchatalbachev, Svetlin; Chakraborty, Trinad; Meinhardt, Andreas; Bhushan, Sudhanshu

    2016-04-01

    Urinary tract infections caused by uropathogenic Escherichia coli (UPEC) pathovars belong to the most frequent infections in human. It is well established that UPEC can subvert innate immune responses, but the role of UPEC in interfering with host cell death pathways is not known. Here, we show that UPEC abrogates activation of the host cell prosurvival protein kinase B signaling pathway, which results in the activation of mammalian forkhead box O (FOXO) transcription factors. Although FOXOs were localized in the nucleus and showed increased DNA-binding activity, no change in the expression levels of FOXO target genes were observed. UPEC can suppress BIM expression induced by LY249002, which results in attenuation of caspase 3 activation and blockage of apoptosis. Mechanistically, BIM expression appears to be epigenetically silenced by a decrease in histone 4 acetylation at the BIM promoter site. Taken together, these results suggest that UPEC can epigenetically silence BIM expression, a molecular switch that prevents apoptosis.

  10. Reef fishes can recognize bleached habitat during settlement: sea anemone bleaching alters anemonefish host selection.

    Science.gov (United States)

    Scott, Anna; Dixson, Danielle L

    2016-05-25

    Understanding how bleaching impacts the settlement of symbiotic habitat specialists and whether there is flexibility in settlement choices with regard to habitat quality is essential given our changing climate. We used five anemonefishes (Amphiprion clarkii, Amphiprion latezonatus, Amphiprion ocellaris, Amphiprion percula and Premnas biaculeatus) and three host sea anemones (Entacmaea quadricolor, Heteractis crispa and Heteractis magnifica) in paired-choice flume experiments to determine whether habitat naive juveniles have the olfactory capabilities to distinguish between unbleached and bleached hosts, and how this may affect settlement decisions. All anemonefishes were able to distinguish between bleached and unbleached hosts, and responded only to chemical cues from species-specific host anemones irrespective of health status, indicating a lack of flexibility in host use. While bleached hosts were selected as habitat, this occurred only when unbleached options were unavailable, with the exception of A. latezonatus, which showed strong preferences for H. crispa regardless of health. This study highlights the potential deleterious indirect impacts of declining habitat quality during larval settlement in habitat specialists, which could be important in the field, given that bleaching events are becoming increasingly common. PMID:27226472

  11. Impact of Adenovirus infection in host cell metabolism evaluated by (1)H-NMR spectroscopy.

    Science.gov (United States)

    Silva, Ana Carina; P Teixeira, Ana; M Alves, Paula

    2016-08-10

    Adenovirus-based vectors are powerful vehicles for gene transfer applications in vaccination and gene therapy. Although highly exploited in the clinical setting, key aspects of the adenovirus biology are still not well understood, in particular the subversion of host cell metabolism during viral infection and replication. The aim of this work was to gain insights on the metabolism of two human cell lines (HEK293 and an amniocyte-derived cell line, 1G3) after infection with an adenovirus serotype 5 vector (AdV5). In order to profile metabolic alterations, we used (1)H-NMR spectroscopy, which allowed the quantification of 35 metabolites in cell culture supernatants with low sample preparation and in a relatively short time. Significant differences between both cell lines in non-infected cultures were identified, namely in glutamine and acetate metabolism, as well as by-product secretion. The main response to AdV5 infection was an increase in glucose consumption and lactate production rates. Moreover, cultures performed with or without glutamine supplementation confirmed the exhaustion of this amino acid as one of the main causes of lower AdV5 production at high cell densities (10- and 1.5-fold less specific yields in HEK293 and 1G3 cells, respectively), and highlighted different degrees of glutamine dependency of adenovirus replication in each cell line. The observed metabolic alterations associated with AdV5 infection and specificity of the host cell line can be useful for targeted bioprocess optimization. PMID:27215342

  12. Tetracycline regulator expression alters the transcriptional program of mammalian cells

    OpenAIRE

    Hackl, Hubert; Rommer, Anna; Konrad, Torsten A; Nassimbeni, Christine; Wieser, Rotraud

    2010-01-01

    Tetracycline regulated ectopic gene expression is a widely used tool to study gene function. However, the tetracycline regulator (tetR) itself has been reported to cause certain phenotypic changes in mammalian cells. We, therefore, asked whether human myeloid U937 cells expressing the tetR in an autoregulated manner would exhibit alterations in gene expression upon removal of tetracycline.

  13. Variation in RNA virus mutation rates across host cells.

    Directory of Open Access Journals (Sweden)

    Marine Combe

    2014-01-01

    Full Text Available It is well established that RNA viruses exhibit higher rates of spontaneous mutation than DNA viruses and microorganisms. However, their mutation rates vary amply, from 10(-6 to 10(-4 substitutions per nucleotide per round of copying (s/n/r and the causes of this variability remain poorly understood. In addition to differences in intrinsic fidelity or error correction capability, viral mutation rates may be dependent on host factors. Here, we assessed the effect of the cellular environment on the rate of spontaneous mutation of the vesicular stomatitis virus (VSV, which has a broad host range and cell tropism. Luria-Delbrück fluctuation tests and sequencing showed that VSV mutated similarly in baby hamster kidney, murine embryonic fibroblasts, colon cancer, and neuroblastoma cells (approx. 10(-5 s/n/r. Cell immortalization through p53 inactivation and oxygen levels (1-21% did not have a significant impact on viral replication fidelity. This shows that previously published mutation rates can be considered reliable despite being based on a narrow and artificial set of laboratory conditions. Interestingly, we also found that VSV mutated approximately four times more slowly in various insect cells compared with mammalian cells. This may contribute to explaining the relatively slow evolution of VSV and other arthropod-borne viruses in nature.

  14. Global impact of Salmonella type III secretion effector SteA on host cells

    Energy Technology Data Exchange (ETDEWEB)

    Cardenal-Muñoz, Elena, E-mail: e_cardenal@us.es; Gutiérrez, Gabriel, E-mail: ggpozo@us.es; Ramos-Morales, Francisco, E-mail: framos@us.es

    2014-07-11

    Highlights: • We analyzed HeLa cells transcriptome in response to Salmonella SteA. • Significant differential expression was detected for 58 human genes. • They are involved in ECM organization and regulation of some signaling pathways. • Cell death, cell adhesion and cell migration were decreased in SteA-expressing cells. • These results contribute to understand the role of SteA during infections. - Abstract: Salmonella enterica is a Gram-negative bacterium that causes gastroenteritis, bacteremia and typhoid fever in several animal species including humans. Its virulence is greatly dependent on two type III secretion systems, encoded in pathogenicity islands 1 and 2. These systems translocate proteins called effectors into eukaryotic host cell. Effectors interfere with host signal transduction pathways to allow the internalization of pathogens and their survival and proliferation inside vacuoles. SteA is one of the few Salmonella effectors that are substrates of both type III secretion systems. Here, we used gene arrays and bioinformatics analysis to study the genetic response of human epithelial cells to SteA. We found that constitutive synthesis of SteA in HeLa cells leads to induction of genes related to extracellular matrix organization and regulation of cell proliferation and serine/threonine kinase signaling pathways. SteA also causes repression of genes related to immune processes and regulation of purine nucleotide synthesis and pathway-restricted SMAD protein phosphorylation. In addition, a cell biology approach revealed that epithelial cells expressing steA show altered cell morphology, and decreased cytotoxicity, cell–cell adhesion and migration.

  15. Phytoplasma adapt to the diverse environments of their plant and insect hosts by altering gene expression

    DEFF Research Database (Denmark)

    Makarova, Olga; MacLean, Allyson M.; Nicolaisen, Mogens

    2015-01-01

    Phytoplasmas are intracellular insect-transmitted phytopathogenic bacteria with small genomes. To understand how Aster Yellows phytoplasma strain witches' broom (AY-WB) adapts to their hosts, we performed qRT-PCR analysis of 179 in silico functionally annotated AY-WB genes that are likely to have...... a role in host adaptation. 74 genes were up-regulated in insects and included genes involved in stress response, phospholipid synthesis, malate and pyruvate metabolism, hemolysin and transporter genes, multiple copies of thymidylate kinase, sigma factor and Zn-proteases genes. In plants, 34 genes...

  16. Alterations in the nuclear proteome of HIV-1 infected T-cells

    Energy Technology Data Exchange (ETDEWEB)

    DeBoer, Jason [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Jagadish, Teena; Haverland, Nicole A. [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Ciborowski, Pawel [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States)

    2014-11-15

    Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines.

  17. Human borna disease virus infection impacts host proteome and histone lysine acetylation in human oligodendroglia cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xia [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Department of Neurology, The Fifth People' s Hospital of Shanghai, School of Medicine, Fudan University, Shanghai, 200240 (China); Zhao, Libo [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Department of Neurology, The Third People' s Hospital of Chongqing, 400014 (China); Yang, Yongtao [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); Bode, Liv [Bornavirus Research Group affiliated to the Free University of Berlin, Berlin (Germany); Huang, Hua [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); Liu, Chengyu [Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); Huang, Rongzhong [Department of Rehabilitative Medicine, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010 (China); Zhang, Liang [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016 (China); Chongqing Key Laboratory of Neurobiology, Chongqing Medical University, Chongqing, 400016 (China); Institute of Neuroscience, Chongqing Medical University, Chongqing, 400016 (China); and others

    2014-09-15

    Background: Borna disease virus (BDV) replicates in the nucleus and establishes persistent infections in mammalian hosts. A human BDV strain was used to address the first time, how BDV infection impacts the proteome and histone lysine acetylation (Kac) of human oligodendroglial (OL) cells, thus allowing a better understanding of infection-driven pathophysiology in vitro. Methods: Proteome and histone lysine acetylation were profiled through stable isotope labeling for cell culture (SILAC)-based quantitative proteomics. The quantifiable proteome was annotated using bioinformatics. Histone acetylation changes were validated by biochemistry assays. Results: Post BDV infection, 4383 quantifiable differential proteins were identified and functionally annotated to metabolism pathways, immune response, DNA replication, DNA repair, and transcriptional regulation. Sixteen of the thirty identified Kac sites in core histones presented altered acetylation levels post infection. Conclusions: BDV infection using a human strain impacted the whole proteome and histone lysine acetylation in OL cells. - Highlights: • A human strain of BDV (BDV Hu-H1) was used to infect human oligodendroglial cells (OL cells). • This study is the first to reveal the host proteomic and histone Kac profiles in BDV-infected OL cells. • BDV infection affected the expression of many transcription factors and several HATs and HDACs.

  18. Human borna disease virus infection impacts host proteome and histone lysine acetylation in human oligodendroglia cells

    International Nuclear Information System (INIS)

    Background: Borna disease virus (BDV) replicates in the nucleus and establishes persistent infections in mammalian hosts. A human BDV strain was used to address the first time, how BDV infection impacts the proteome and histone lysine acetylation (Kac) of human oligodendroglial (OL) cells, thus allowing a better understanding of infection-driven pathophysiology in vitro. Methods: Proteome and histone lysine acetylation were profiled through stable isotope labeling for cell culture (SILAC)-based quantitative proteomics. The quantifiable proteome was annotated using bioinformatics. Histone acetylation changes were validated by biochemistry assays. Results: Post BDV infection, 4383 quantifiable differential proteins were identified and functionally annotated to metabolism pathways, immune response, DNA replication, DNA repair, and transcriptional regulation. Sixteen of the thirty identified Kac sites in core histones presented altered acetylation levels post infection. Conclusions: BDV infection using a human strain impacted the whole proteome and histone lysine acetylation in OL cells. - Highlights: • A human strain of BDV (BDV Hu-H1) was used to infect human oligodendroglial cells (OL cells). • This study is the first to reveal the host proteomic and histone Kac profiles in BDV-infected OL cells. • BDV infection affected the expression of many transcription factors and several HATs and HDACs

  19. RNAi screen reveals host cell kinases specifically involved in Listeria monocytogenes spread from cell to cell.

    Directory of Open Access Journals (Sweden)

    Ryan Chong

    Full Text Available Intracellular bacterial pathogens, such as Listeria monocytogenes and Rickettsia conorii display actin-based motility in the cytosol of infected cells and spread from cell to cell through the formation of membrane protrusions at the cell cortex. Whereas the mechanisms supporting cytosolic actin-based motility are fairly well understood, it is unclear whether specific host factors may be required for supporting the formation and resolution of membrane protrusions. To address this gap in knowledge, we have developed high-throughput fluorescence microscopy and computer-assisted image analysis procedures to quantify pathogen spread in human epithelial cells. We used the approach to screen a siRNA library covering the human kinome and identified 7 candidate kinases whose depletion led to severe spreading defects in cells infected with L. monocytogenes. We conducted systematic validation procedures with redundant silencing reagents and confirmed the involvement of the serine/threonine kinases, CSNK1A1 and CSNK2B. We conducted secondary assays showing that, in contrast with the situation observed in CSNK2B-depleted cells, L. monocytogenes formed wild-type cytosolic tails and displayed wild-type actin-based motility in the cytosol of CSNK1A1-depleted cells. Furthermore, we developed a protrusion formation assay and showed that the spreading defect observed in CSNK1A1-depleted cells correlated with the formation of protrusion that did not resolve into double-membrane vacuoles. Moreover, we developed sending and receiving cell-specific RNAi procedures and showed that CSNK1A was required in the sending cells, but was dispensable in the receiving cells, for protrusion resolution. Finally, we showed that the observed defects were specific to Listeria monocytogenes, as Rickettsia conorii displayed wild-type cell-to-cell spread in CSNK1A1- and CSNK2B-depleted cells. We conclude that, in addition to the specific host factors supporting cytosolic actin

  20. Radiation-induced motility alterations in medulloblastoma cells

    OpenAIRE

    Rieken, Stefan; Rieber, Juliane; Brons, Stephan; Habermehl, Daniel; Rief, Harald; Orschiedt, Lena; Lindel, Katja; Klaus J. Weber; Debus, Jürgen; Combs, Stephanie E

    2015-01-01

    Photon irradiation has been repeatedly suspected of increasing tumor cell motility and promoting locoregional recurrence of disease. This study was set up to analyse possible mechanisms underlying the potentially radiation-altered motility in medulloblastoma cells. Medulloblastoma cell lines D425 and Med8A were analyzed in migration and adhesion experiments with and without photon and carbon ion irradiation. Expression of integrins was determined by quantitative FACS analysis. Matrix metallop...

  1. Nematomorph parasites indirectly alter the food web and ecosystem function of streams through behavioural manipulation of their cricket hosts.

    Science.gov (United States)

    Sato, T.; Egusa, T.; Fukushima, K.; Oda, T.; Ohte, N.; Tokuchi, Naoko; Watanabe, Katsutoshi; Kanaiwa, Minoru; Murakami, Isaya; Lafferty, Kevin D.

    2012-01-01

    Nematomorph parasites manipulate crickets to enter streams where the parasites reproduce. These manipulated crickets become a substantial food subsidy for stream fishes. We used a field experiment to investigate how this subsidy affects the stream community and ecosystem function. When crickets were available, predatory fish ate fewer benthic invertebrates. The resulting release of the benthic invertebrate community from fish predation indirectly decreased the biomass of benthic algae and slightly increased leaf break-down rate. This is the first experimental demonstration that host manipulation by a parasite can reorganise a community and alter ecosystem function. Nematomorphs are common, and many other parasites have dramatic effects on host phenotypes, suggesting that similar effects of parasites on ecosystems might be widespread.

  2. Metabolic alterations in cancer cells and therapeutic implications

    Institute of Scientific and Technical Information of China (English)

    Naima Hammoudi; Kausar Begam Riaz Ahmed; Celia Garcia-Prieto; Peng Huang

    2011-01-01

    Cancer metabolism has emerged as an important area of research in recent years. Elucidation of the metabolic differences between cancer and normal cells and the underlying mechanisms will not only advance our understanding of fundamental cancer cell biology but also provide an important basis for the development of new therapeutic strategies and novel compounds to selectively eliminate cancer cells by targeting their unique metabolism. This article reviews several important metabolic alterations in cancer cells, with an emphasis on increased aerobic glycolysis (the Warburg effect) and glutamine addiction, and discusses the mechanisms that may contribute to such metabolic changes. In addition, metabolic alterations in cancer stem cells, mitochondrial metabolism and its influence on drug sensitivity, and potential therapeutic strategies and agents that target cancer metabolism are also discussed.

  3. Anthelmintic treatment alters the parasite community in a wild mouse host.

    Science.gov (United States)

    Pedersen, Amy B; Antonovics, Janis

    2013-08-23

    Individuals are often co-infected with several parasite species, yet the consequences of drug treatment on the dynamics of parasite communities in wild populations have rarely been measured. Here, we experimentally reduced nematode infection in a wild mouse population and measured the effects on other non-target parasites. A single oral dose of the anthelmintic, ivermectin, significantly reduced nematode infection, but resulted in a reciprocal increase in other gastrointestinal parasites, specifically coccidial protozoans and cestodes. These results highlight the possibility that drug therapy may have unintended consequences for non-target parasites and that host-parasite dynamics cannot always be fully understood in the framework of single host-parasite interactions.

  4. Genome sequence alterations detected upon passage of Burkholderia mallei ATCC 23344 in culture and in mammalian hosts

    Directory of Open Access Journals (Sweden)

    Yu Yan

    2006-09-01

    Full Text Available Abstract Background More than 12,000 simple sequence repeats (SSRs have been identified in the genome of Burkholderia mallei ATCC 23344. As a demonstrated mechanism of phase variation in other pathogenic bacteria, these may function as mutable loci leading to altered protein expression or structure variation. To determine if such alterations are occurring in vivo, the genomes of various single-colony passaged B. mallei ATCC 23344 isolates, one from each source, were sequenced from culture, a mouse, a horse, and two isolates from a single human patient, and the sequence compared to the published B. mallei ATCC 23344 genome sequence. Results Forty-nine insertions and deletions (indels were detected at SSRs in the five passaged strains, a majority of which (67.3% were located within noncoding areas, suggesting that such regions are more tolerant of sequence alterations. Expression profiling of the two human passaged isolates compared to the strain before passage revealed alterations in the mRNA levels of multiple genes when grown in culture. Conclusion These data support the notion that genome variability upon passage is a feature of B. mallei ATCC23344, and that within a host B. mallei generates a diverse population of clones that accumulate genome sequence variation at SSR and other loci.

  5. Climate change may alter breeding ground distributions of eastern migratory monarchs (Danaus plexippus via range expansion of Asclepias host plants.

    Directory of Open Access Journals (Sweden)

    Nathan P Lemoine

    Full Text Available Climate change can profoundly alter species' distributions due to changes in temperature, precipitation, or seasonality. Migratory monarch butterflies (Danaus plexippus may be particularly susceptible to climate-driven changes in host plant abundance or reduced overwintering habitat. For example, climate change may significantly reduce the availability of overwintering habitat by restricting the amount of area with suitable microclimate conditions. However, potential effects of climate change on monarch northward migrations remain largely unknown, particularly with respect to their milkweed (Asclepias spp. host plants. Given that monarchs largely depend on the genus Asclepias as larval host plants, the effects of climate change on monarch northward migrations will most likely be mediated by climate change effects on Asclepias. Here, I used MaxEnt species distribution modeling to assess potential changes in Asclepias and monarch distributions under moderate and severe climate change scenarios. First, Asclepias distributions were projected to extend northward throughout much of Canada despite considerable variability in the environmental drivers of each individual species. Second, Asclepias distributions were an important predictor of current monarch distributions, indicating that monarchs may be constrained as much by the availability of Asclepias host plants as environmental variables per se. Accordingly, modeling future distributions of monarchs, and indeed any tightly coupled plant-insect system, should incorporate the effects of climate change on host plant distributions. Finally, MaxEnt predictions of Asclepias and monarch distributions were remarkably consistent among general circulation models. Nearly all models predicted that the current monarch summer breeding range will become slightly less suitable for Asclepias and monarchs in the future. Asclepias, and consequently monarchs, should therefore undergo expanded northern range limits in

  6. Climate change may alter breeding ground distributions of eastern migratory monarchs (Danaus plexippus) via range expansion of Asclepias host plants.

    Science.gov (United States)

    Lemoine, Nathan P

    2015-01-01

    Climate change can profoundly alter species' distributions due to changes in temperature, precipitation, or seasonality. Migratory monarch butterflies (Danaus plexippus) may be particularly susceptible to climate-driven changes in host plant abundance or reduced overwintering habitat. For example, climate change may significantly reduce the availability of overwintering habitat by restricting the amount of area with suitable microclimate conditions. However, potential effects of climate change on monarch northward migrations remain largely unknown, particularly with respect to their milkweed (Asclepias spp.) host plants. Given that monarchs largely depend on the genus Asclepias as larval host plants, the effects of climate change on monarch northward migrations will most likely be mediated by climate change effects on Asclepias. Here, I used MaxEnt species distribution modeling to assess potential changes in Asclepias and monarch distributions under moderate and severe climate change scenarios. First, Asclepias distributions were projected to extend northward throughout much of Canada despite considerable variability in the environmental drivers of each individual species. Second, Asclepias distributions were an important predictor of current monarch distributions, indicating that monarchs may be constrained as much by the availability of Asclepias host plants as environmental variables per se. Accordingly, modeling future distributions of monarchs, and indeed any tightly coupled plant-insect system, should incorporate the effects of climate change on host plant distributions. Finally, MaxEnt predictions of Asclepias and monarch distributions were remarkably consistent among general circulation models. Nearly all models predicted that the current monarch summer breeding range will become slightly less suitable for Asclepias and monarchs in the future. Asclepias, and consequently monarchs, should therefore undergo expanded northern range limits in summer months

  7. Role of curcumin-dependent modulation of tumor microenvironment of a murine T cell lymphoma in altered regulation of tumor cell survival

    International Nuclear Information System (INIS)

    Using a murine model of a T cell lymphoma, in the present study, we report that tumor growth retarding action of curcumin involves modulation of some crucial parameters of tumor microenvironment regulating tumor progression. Curcumin-administration to tumor-bearing host caused an altered pH regulation in tumor cells associated with alteration in expression of cell survival and apoptosis regulatory proteins and genes. Nevertheless, an alteration was also observed in biophysical parameters of tumor microenvironment responsible for modulation of tumor growth pertaining to hypoxia, tumor acidosis, and glucose metabolism. The study thus sheds new light with respect to the antineoplastic action of curcumin against a tumor-bearing host with progressively growing tumor of hematological origin. This will help in optimizing application of the drug and anticancer research and therapy. - Graphical Abstract: Display Omitted

  8. Staphylococcus aureus-induced G2/M phase transition delay in host epithelial cells increases bacterial infective efficiency.

    Science.gov (United States)

    Alekseeva, Ludmila; Rault, Lucie; Almeida, Sintia; Legembre, Patrick; Edmond, Valérie; Azevedo, Vasco; Miyoshi, Anderson; Even, Sergine; Taieb, Frédéric; Arlot-Bonnemains, Yannick; Le Loir, Yves; Berkova, Nadia

    2013-01-01

    Staphylococcus aureus is a highly versatile, opportunistic pathogen and the etiological agent of a wide range of infections in humans and warm-blooded animals. The epithelial surface is its principal site of colonization and infection. In this work, we investigated the cytopathic effect of S. aureus strains from human and animal origins and their ability to affect the host cell cycle in human HeLa and bovine MAC-T epithelial cell lines. S. aureus invasion slowed down cell proliferation and induced a cytopathic effect, resulting in the enlargement of host cells. A dramatic decrease in the number of mitotic cells was observed in the infected cultures. Flow cytometry analysis revealed an S. aureus-induced delay in the G2/M phase transition in synchronous HeLa cells. This delay required the presence of live S. aureus since the addition of the heat-killed bacteria did not alter the cell cycle. The results of Western blot experiments showed that the G2/M transition delay was associated with the accumulation of inactive cyclin-dependent kinase Cdk1, a key inducer of mitosis entry, and with the accumulation of unphosphorylated histone H3, which was correlated with a reduction of the mitotic cell number. Analysis of S. aureus proliferation in asynchronous, G1- and G2-phase-enriched HeLa cells showed that the G2 phase was preferential for bacterial infective efficiency, suggesting that the G2 phase delay may be used by S. aureus for propagation within the host. Taken together, our results divulge the potential of S. aureus in the subversion of key cellular processes such as cell cycle progression, and shed light on the biological significance of S. aureus-induced host cell cycle alteration.

  9. Staphylococcus aureus-induced G2/M phase transition delay in host epithelial cells increases bacterial infective efficiency.

    Directory of Open Access Journals (Sweden)

    Ludmila Alekseeva

    Full Text Available Staphylococcus aureus is a highly versatile, opportunistic pathogen and the etiological agent of a wide range of infections in humans and warm-blooded animals. The epithelial surface is its principal site of colonization and infection. In this work, we investigated the cytopathic effect of S. aureus strains from human and animal origins and their ability to affect the host cell cycle in human HeLa and bovine MAC-T epithelial cell lines. S. aureus invasion slowed down cell proliferation and induced a cytopathic effect, resulting in the enlargement of host cells. A dramatic decrease in the number of mitotic cells was observed in the infected cultures. Flow cytometry analysis revealed an S. aureus-induced delay in the G2/M phase transition in synchronous HeLa cells. This delay required the presence of live S. aureus since the addition of the heat-killed bacteria did not alter the cell cycle. The results of Western blot experiments showed that the G2/M transition delay was associated with the accumulation of inactive cyclin-dependent kinase Cdk1, a key inducer of mitosis entry, and with the accumulation of unphosphorylated histone H3, which was correlated with a reduction of the mitotic cell number. Analysis of S. aureus proliferation in asynchronous, G1- and G2-phase-enriched HeLa cells showed that the G2 phase was preferential for bacterial infective efficiency, suggesting that the G2 phase delay may be used by S. aureus for propagation within the host. Taken together, our results divulge the potential of S. aureus in the subversion of key cellular processes such as cell cycle progression, and shed light on the biological significance of S. aureus-induced host cell cycle alteration.

  10. Improved metabolic health alters host metabolism in parallel with changes in systemic xeno-metabolites of gut origin.

    Directory of Open Access Journals (Sweden)

    Caitlin Campbell

    Full Text Available Novel plasma metabolite patterns reflective of improved metabolic health (insulin sensitivity, fitness, reduced body weight were identified before and after a 14-17 wk weight loss and exercise intervention in sedentary, obese insulin-resistant women. To control for potential confounding effects of diet- or microbiome-derived molecules on the systemic metabolome, sampling was during a tightly-controlled feeding test week paradigm. Pairwise and multivariate analysis revealed intervention- and insulin-sensitivity associated: (1 Changes in plasma xeno-metabolites ("non-self" metabolites of dietary or gut microbial origin following an oral glucose tolerance test (e.g. higher post-OGTT propane-1,2,3-tricarboxylate [tricarballylic acid] or in the overnight-fasted state (e.g., lower γ-tocopherol; (2 Increased indices of saturated very long chain fatty acid elongation capacity; (3 Increased post-OGTT α-ketoglutaric acid (α-KG, fasting α-KG inversely correlated with Matsuda index, and altered patterns of malate, pyruvate and glutamine hypothesized to stem from improved mitochondrial efficiency and more robust oxidation of glucose. The results support a working model in which improved metabolic health modifies host metabolism in parallel with altering systemic exposure to xeno-metabolites. This highlights that interpretations regarding the origins of peripheral blood or urinary "signatures" of insulin resistance and metabolic health must consider the potentially important contribution of gut-derived metabolites toward the host's metabolome.

  11. Improved metabolic health alters host metabolism in parallel with changes in systemic xeno-metabolites of gut origin.

    Science.gov (United States)

    Campbell, Caitlin; Grapov, Dmitry; Fiehn, Oliver; Chandler, Carol J; Burnett, Dustin J; Souza, Elaine C; Casazza, Gretchen A; Gustafson, Mary B; Keim, Nancy L; Newman, John W; Hunter, Gary R; Fernandez, Jose R; Garvey, W Timothy; Harper, Mary-Ellen; Hoppel, Charles L; Meissen, John K; Take, Kohei; Adams, Sean H

    2014-01-01

    Novel plasma metabolite patterns reflective of improved metabolic health (insulin sensitivity, fitness, reduced body weight) were identified before and after a 14-17 wk weight loss and exercise intervention in sedentary, obese insulin-resistant women. To control for potential confounding effects of diet- or microbiome-derived molecules on the systemic metabolome, sampling was during a tightly-controlled feeding test week paradigm. Pairwise and multivariate analysis revealed intervention- and insulin-sensitivity associated: (1) Changes in plasma xeno-metabolites ("non-self" metabolites of dietary or gut microbial origin) following an oral glucose tolerance test (e.g. higher post-OGTT propane-1,2,3-tricarboxylate [tricarballylic acid]) or in the overnight-fasted state (e.g., lower γ-tocopherol); (2) Increased indices of saturated very long chain fatty acid elongation capacity; (3) Increased post-OGTT α-ketoglutaric acid (α-KG), fasting α-KG inversely correlated with Matsuda index, and altered patterns of malate, pyruvate and glutamine hypothesized to stem from improved mitochondrial efficiency and more robust oxidation of glucose. The results support a working model in which improved metabolic health modifies host metabolism in parallel with altering systemic exposure to xeno-metabolites. This highlights that interpretations regarding the origins of peripheral blood or urinary "signatures" of insulin resistance and metabolic health must consider the potentially important contribution of gut-derived metabolites toward the host's metabolome. PMID:24416208

  12. Mycobacterium tuberculosis co-operonic PE32/PPE65 proteins alter host immune responses by hampering Th1 response

    Directory of Open Access Journals (Sweden)

    Mohd eKhubaib

    2016-05-01

    Full Text Available PE/PPE genes, present in cluster with ESAT-6 like genes, are suspected to have a role in antigenic variation and virulence of Mycobacterium tuberculosis. Their roles in immune evasion and immune modulation of host are also well documented. We present evidence that PE32/PPE65 present within the RD8 region are co-operonic, co-transcribed and co-translated, and play role in modulating host immune responses. Experiments with macrophage cell lines revealed that this protein complex suppresses pro-inflammatory cytokines such as TNF-α and IL-6 whereas also inducing high expression of anti-inflammatory IL-10. Immunization of mice with these recombinant proteins dampens an effective Th1 response as evident from reduced frequency of IFN-g and IL-2 producing CD4+ and CD8+ T cells. IgG sub-typing from serum of immunized mice revealed high levels of IgG1 when compared with IgG2a and IgG2b. Further IgG1/IgG2a ratio clearly demonstrated that the protein complex manipulates the host immune response favourable to the pathogen. Our results demonstrate that the co-transcribed and co-translated PE32 and PPE65 antigens are involved specifically in modulating anti-mycobacterial host immune response by hampering Th1 response.

  13. Radiation-induced motility alterations in medulloblastoma cells.

    Science.gov (United States)

    Rieken, Stefan; Rieber, Juliane; Brons, Stephan; Habermehl, Daniel; Rief, Harald; Orschiedt, Lena; Lindel, Katja; Weber, Klaus J; Debus, Jürgen; Combs, Stephanie E

    2015-05-01

    Photon irradiation has been repeatedly suspected of increasing tumor cell motility and promoting locoregional recurrence of disease. This study was set up to analyse possible mechanisms underlying the potentially radiation-altered motility in medulloblastoma cells. Medulloblastoma cell lines D425 and Med8A were analyzed in migration and adhesion experiments with and without photon and carbon ion irradiation. Expression of integrins was determined by quantitative FACS analysis. Matrix metalloproteinase concentrations within cell culture supernatants were investigated by enzyme-linked immunosorbent assay (ELISA). Statistical analysis was performed using Student's t-test. Both photon and carbon ion irradiation significantly reduced chemotactic medulloblastoma cell transmigration through 8-μm pore size membranes, while simultaneously increasing adherence to fibronectin- and collagen I- and IV-coated surfaces. Correspondingly, both photon and carbon ion irradiation downregulate soluble MMP9 concentrations, while upregulating cell surface expression of proadhesive extracellular matrix protein-binding integrin α5. The observed phenotype of radiation-altered motility is more pronounced following carbon ion than photon irradiation. Both photon and (even more so) carbon ion irradiation are effective in inhibiting medulloblastoma cell migration through downregulation of matrix metalloproteinase 9 and upregulation of proadhesive cell surface integrin α5, which lead to increased cell adherence to extracellular matrix proteins. PMID:25736470

  14. Structural determinants of host defense peptides for antimicrobial activity and target cell selectivity.

    Science.gov (United States)

    Takahashi, Daisuke; Shukla, Sanjeev K; Prakash, Om; Zhang, Guolong

    2010-09-01

    Antimicrobial host defense peptides (HDPs) are a critical component of the innate immunity with microbicidal, endotoxin-neutralizing, and immunostimulatory properties. HDPs kill bacteria primarily through non-specific membrane lysis, therefore with a less likelihood of provoking resistance. Extensive structure-activity relationship studies with a number of HDPs have revealed that net charge, amphipathicity, hydrophobicity, and structural propensity are among the most important physicochemical and structural parameters that dictate their ability to interact with and disrupt membranes. A delicate balance among these factors, rather than a mere alteration of a single factor, is critically important for HDPs to ensure the antimicrobial potency and target cell selectivity. With a better understanding of the structural determinants of HDPs for their membrane-lytic activities, it is expected that novel HDP-based antimicrobials with minimum toxicity to eukaryotic cells can be developed for resistant infections, which have become a global public health crisis.

  15. How stem cells speak with host immune cells in inflammatory brain diseases.

    Science.gov (United States)

    Pluchino, Stefano; Cossetti, Chiara

    2013-09-01

    Advances in stem cell biology have raised great expectations that diseases and injuries of the central nervous system (CNS) may be ameliorated by the development of non-hematopoietic stem cell medicines. Yet, the application of adult stem cells as CNS therapeutics is challenging and the interpretation of some of the outcomes ambiguous. In fact, the initial idea that stem cell transplants work only via structural cell replacement has been challenged by the observation of consistent cellular signaling between the graft and the host. Cellular signaling is the foundation of coordinated actions and flexible responses, and arises via networks of exchanging and interacting molecules that transmit patterns of information between cells. Sustained stem cell graft-to-host communication leads to remarkable trophic effects on endogenous brain cells and beneficial modulatory actions on innate and adaptive immune responses in vivo, ultimately promoting the healing of the injured CNS. Among a number of adult stem cell types, mesenchymal stem cells (MSCs) and neural stem/precursor cells (NPCs) are being extensively investigated for their ability to signal to the immune system upon transplantation in experimental CNS diseases. Here, we focus on the main cellular signaling pathways that grafted MSCs and NPCs use to establish a therapeutically relevant cross talk with host immune cells, while examining the role of inflammation in regulating some of the bidirectionality of these communications. We propose that the identification of the players involved in stem cell signaling might contribute to the development of innovative, high clinical impact therapeutics for inflammatory CNS diseases.

  16. A microfluidic cell-trapping device for single-cell tracking of host-microbe interactions.

    Science.gov (United States)

    Delincé, Matthieu J; Bureau, Jean-Baptiste; López-Jiménez, Ana Teresa; Cosson, Pierre; Soldati, Thierry; McKinney, John D

    2016-08-16

    The impact of cellular individuality on host-microbe interactions is increasingly appreciated but studying the temporal dynamics of single-cell behavior in this context remains technically challenging. Here we present a microfluidic platform, InfectChip, to trap motile infected cells for high-resolution time-lapse microscopy. This approach allows the direct visualization of all stages of infection, from bacterial uptake to death of the bacterium or host cell, over extended periods of time. We demonstrate the utility of this approach by co-culturing an established host-cell model, Dictyostelium discoideum, with the extracellular pathogen Klebsiella pneumoniae or the intracellular pathogen Mycobacterium marinum. We show that the outcome of such infections is surprisingly heterogeneous, ranging from abortive infection to death of the bacterium or host cell. InfectChip thus provides a simple method to dissect the time-course of host-microbe interactions at the single-cell level, yielding new insights that could not be gleaned from conventional population-based measurements. PMID:27425421

  17. Emerging functions as host cell factors - an encyclopedia of annexin-pathogen interactions.

    Science.gov (United States)

    Kuehnl, Alexander; Musiol, Agnes; Raabe, Carsten A; Rescher, Ursula

    2016-10-01

    Emerging infectious diseases and drug-resistant infectious agents call for the development of innovative antimicrobial strategies. With pathogenicity now considered to arise from the complex and bi-directional interplay between a microbe and the host, host cell factor targeting has emerged as a promising approach that might overcome the limitations of classical antimicrobial drug development and could open up novel and efficient therapeutic strategies. Interaction with and modulation of host cell membranes is a recurrent theme in the host-microbe relationship. In this review, we provide an overview of what is currently known about the role of the Ca2+ dependent, membrane-binding annexin protein family in pathogen-host interactions, and discuss their emerging functions as host cell derived auxiliary proteins in microbe-host interactions and host cell targets.

  18. Cell elasticity with altered cytoskeletal architectures across multiple cell types.

    Science.gov (United States)

    Grady, Martha E; Composto, Russell J; Eckmann, David M

    2016-08-01

    The cytoskeleton is primarily responsible for providing structural support, localization and transport of organelles, and intracellular trafficking. The structural support is supplied by actin filaments, microtubules, and intermediate filaments, which contribute to overall cell elasticity to varying degrees. We evaluate cell elasticity in five different cell types with drug-induced cytoskeletal derangements to probe how actin filaments and microtubules contribute to cell elasticity and whether it is conserved across cell type. Specifically, we measure elastic stiffness in primary chondrocytes, fibroblasts, endothelial cells (HUVEC), hepatocellular carcinoma cells (HUH-7), and fibrosarcoma cells (HT 1080) subjected to two cytoskeletal destabilizers: cytochalasin D and nocodazole, which disrupt actin and microtubule polymerization, respectively. Elastic stiffness is measured by atomic force microscopy (AFM) and the disruption of the cytoskeleton is confirmed using fluorescence microscopy. The two cancer cell lines showed significantly reduced elastic moduli values (~0.5kPa) when compared to the three healthy cell lines (~2kPa). Non-cancer cells whose actin filaments were disrupted using cytochalasin D showed a decrease of 60-80% in moduli values compared to untreated cells of the same origin, whereas the nocodazole-treated cells showed no change in elasticity. Overall, we demonstrate actin filaments contribute more to elastic stiffness than microtubules but this result is cell type dependent. Cancer cells behaved differently, exhibiting increased stiffness as well as stiffness variability when subjected to nocodazole. We show that disruption of microtubule dynamics affects cancer cell elasticity, suggesting therapeutic drugs targeting microtubules be monitored for significant elastic changes. PMID:26874250

  19. The transforming parasite Theileria co-opts host cell mitotic and central spindles to persist in continuously dividing cells.

    Directory of Open Access Journals (Sweden)

    Conrad von Schubert

    Full Text Available The protozoan parasite Theileria inhabits the host cell cytoplasm and possesses the unique capacity to transform the cells it infects, inducing continuous proliferation and protection against apoptosis. The transforming schizont is a multinucleated syncytium that resides free in the host cell cytoplasm and is strictly intracellular. To maintain transformation, it is crucial that this syncytium is divided over the two daughter cells at each host cell cytokinesis. This process was dissected using different cell cycle synchronization methods in combination with the targeted application of specific inhibitors. We found that Theileria schizonts associate with newly formed host cell microtubules that emanate from the spindle poles, positioning the parasite at the equatorial region of the mitotic cell where host cell chromosomes assemble during metaphase. During anaphase, the schizont interacts closely with host cell central spindle. As part of this process, the schizont recruits a host cell mitotic kinase, Polo-like kinase 1, and we established that parasite association with host cell central spindles requires Polo-like kinase 1 catalytic activity. Blocking the interaction between the schizont and astral as well as central spindle microtubules prevented parasite segregation between the daughter cells during cytokinesis. Our findings provide a striking example of how an intracellular eukaryotic pathogen that evolved ways to induce the uncontrolled proliferation of the cells it infects usurps the host cell mitotic machinery, including Polo-like kinase 1, one of the pivotal mitotic kinases, to ensure its own persistence and survival.

  20. Bacterial colonization of host cells in the absence of cholesterol.

    Directory of Open Access Journals (Sweden)

    Stacey D Gilk

    2013-01-01

    Full Text Available Reports implicating important roles for cholesterol and cholesterol-rich lipid rafts in host-pathogen interactions have largely employed sterol sequestering agents and biosynthesis inhibitors. Because the pleiotropic effects of these compounds can complicate experimental interpretation, we developed a new model system to investigate cholesterol requirements in pathogen infection utilizing DHCR24(-/- mouse embryonic fibroblasts (MEFs. DHCR24(-/- MEFs lack the Δ24 sterol reductase required for the final enzymatic step in cholesterol biosynthesis, and consequently accumulate desmosterol into cellular membranes. Defective lipid raft function by DHCR24(-/- MEFs adapted to growth in cholesterol-free medium was confirmed by showing deficient uptake of cholera-toxin B and impaired signaling by epidermal growth factor. Infection in the absence of cholesterol was then investigated for three intracellular bacterial pathogens: Coxiella burnetii, Salmonella enterica serovar Typhimurium, and Chlamydia trachomatis. Invasion by S. Typhimurium and C. trachomatis was unaltered in DHCR24(-/- MEFs. In contrast, C. burnetii entry was significantly decreased in -cholesterol MEFs, and also in +cholesterol MEFs when lipid raft-associated α(Vβ(3 integrin was blocked, suggesting a role for lipid rafts in C. burnetii uptake. Once internalized, all three pathogens established their respective vacuolar niches and replicated normally. However, the C. burnetii-occupied vacuole within DHCR24(-/- MEFs lacked the CD63-positive material and multilamellar membranes typical of vacuoles formed in wild type cells, indicating cholesterol functions in trafficking of multivesicular bodies to the pathogen vacuole. These data demonstrate that cholesterol is not essential for invasion and intracellular replication by S. Typhimurium and C. trachomatis, but plays a role in C. burnetii-host cell interactions.

  1. Parasitic castration by the digenian trematode Allopodocotyle sp. alters gene expression in the brain of the host mollusc Haliotis asinina.

    Science.gov (United States)

    Rice, Tamika; McGraw, Elizabeth; O'Brien, Elizabeth K; Reverter, Antonio; Jackson, Daniel J; Degnan, Bernard M

    2006-06-26

    Infection of molluscs by digenean trematode parasites typically results in the repression of reproduction -- the so-called parasitic castration. This is known to occur by altering the expression of a range of host neuropeptide genes. Here we analyse the expression levels of 10 members of POU, Pax, Sox and Hox transcription factor gene families, along with genes encoding FMRFamide, prohormone convertase and beta-tubulin, in the brain ganglia of actively reproducing (summer), non-reproducing (winter) and infected Haliotis asinina (a vetigastropod mollusc). A number of the regulatory genes are differentially expressed in parasitised H. asinina, but in only a few cases do expression patterns in infected animals match those occurring in animals where reproduction is normally repressed.

  2. Interaction of KSHV with Host Cell Surface Receptors and Cell Entry

    Directory of Open Access Journals (Sweden)

    Mohanan Valiya Veettil

    2014-10-01

    Full Text Available Virus entry is a complex process characterized by a sequence of events. Since the discovery of KSHV in 1994, tremendous progress has been made in our understanding of KSHV entry into its in vitro target cells. KSHV entry is a complex multistep process involving viral envelope glycoproteins and several cell surface molecules that is utilized by KSHV for its attachment and entry. KSHV has a broad cell tropism and the attachment and receptor engagement on target cells have an important role in determining the cell type-specific mode of entry. KSHV utilizes heparan sulfate, integrins and EphrinA2 molecules as receptors which results in the activation of host cell pre-existing signal pathways that facilitate the subsequent cascade of events resulting in the rapid entry of virus particles, trafficking towards the nucleus followed by viral and host gene expression. KSHV enters human fibroblast cells by dynamin dependant clathrin mediated endocytosis and by dynamin independent macropinocytosis in dermal endothelial cells. Once internalized into endosomes, fusion of the viral envelope with the endosomal membranes in an acidification dependent manner results in the release of capsids which subsequently reaches the nuclear pore vicinity leading to the delivery of viral DNA into the nucleus. In this review, we discuss the principal mechanisms that enable KSHV to interact with the host cell surface receptors as well as the mechanisms that are required to modulate cell signaling machinery for a successful entry.

  3. Altered cytoskeletal structures in transformed cells exhibiting obviously metastatic capabilities

    Institute of Scientific and Technical Information of China (English)

    LINZHONGXIANG; WUBINGQUAN; 等

    1990-01-01

    Cytoskeletal changes in transformed cells (LM-51) eshibiting obviously metastatic capabilities were investigated by utilization of double-fluorescent labelling through combinations of:(1) tubulin indirect immunofluorescence plus Rhodamine-phalloidin staining of F-actins;(2) indirect immunofluorescent staining with α-actinin polyclonal-and vinculin monoclonal antibodies.The LM-51 cells which showed metastatic index of >50% were derived from lung metastasis in nude mice after subcutaneous inoculation of human highly metastatic tumor DNA transfected NIH3T3 cell transformants.The parent NIH3T3 cells exhibited well-organized microtubules,prominent stress fibers and adhesion plaques while their transformants showed remarkable cytoskeletal alterations:(1)reduced microtubules but increased MTOC fluorescence;(2)disrupted stress fibers and fewer adhesion plaques with their protein components redistributed in the cytoplasm;(3)Factin-and α-actinin/vinculin aggregates appeared in the cytoplasm.These aggregates were dot-like,varied in size(0.1-0.4μm) and number,located near the ventral surface of the cells.TPA-induced actin/vinculin bodies were studied too.Indications that actin and α-actinin/vinculin redistribution might be important alterations involved in the expression of metastatic capabilities of LM-51 transformed cells were discussed.

  4. Does vegetation complexity affect host plant chemistry, and thus multitrophic interactions, in a human-altered landscape?

    Science.gov (United States)

    Wäschke, Nicole; Hancock, Christine; Hilker, Monika; Obermaier, Elisabeth; Meiners, Torsten

    2015-09-01

    Anthropogenic land use may shape vegetation composition and affect trophic interactions by altering concentrations of host plant metabolites. Here, we investigated the hypotheses that: (1) plant N and defensive secondary metabolite contents of the herb Plantago lanceolata are affected by land use intensity (LUI) and the surrounding vegetation composition (=plant species richness and P. lanceolata density), and that (2) changes in plant chemistry affect abundances of the herbivorous weevils Mecinus pascuorum and Mecinus labilis, as well as their larval parasitoid Mesopolobus incultus, in the field. We determined plant species richness, P. lanceolata density, and abundances of the herbivores and the parasitoid in 77 grassland plots differing in LUI index in three regions across Germany. We also measured the N and secondary metabolite [the iridoid glycosides (IGs) aucubin and catalpol] contents of P. lanceolata leaves. Mixed-model analysis revealed that: (1) concentrations of leaf IGs were positively correlated with plant species richness; leaf N content was positively correlated with the LUI index. Furthermore: (2) herbivore abundance was not related to IG concentrations, but correlated negatively with leaf N content. Parasitoid abundance correlated positively only with host abundance over the three regions. Structural equation models revealed a positive impact of IG concentrations on parasitoid abundance in one region. We conclude that changes in plant chemistry due to land use and/or vegetation composition may affect higher trophic levels and that the manifestation of these effects may depend on local biotic or abiotic features of the landscape. PMID:25986560

  5. The perfect host: a mouse host embryo facilitating more efficient germ line transmission of genetically modified embryonic stem cells.

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    Robert A Taft

    Full Text Available There is a continual need to improve efficiency in creating precise genetic modifications in mice using embryonic stem cells (ESCs. We describe a novel approach resulting in 100% germline transmission from competent injected ESCs. We developed an F1 mouse host embryo (Perfect Host, PH that selectively ablates its own germ cells via tissue-specific induction of diphtheria toxin. This approach allows competent microinjected ESCs to fully dominate the germline, eliminating competition for this critical niche in the developing and adult animal. This is in contrast to conventional methods, where competition from host germ cells results in offspring derived from host cells and ESCs, necessitating extensive breeding of chimeras and genotyping to identify germline. The germline transmission process is also complicated by variability in the actual number of ESCs that colonize the germline niche and the proportion that are germline competent. To validate the PH approach we used ESC lines derived from 129 F1, BALB/cByJ, and BTBR backgrounds as well as an iPS line. Resulting chimeric males produced 194 offspring, all paternally derived from the introduced stem cells, with no offspring being derived from the host genome. We further tested this approach using eleven genetically modified C57BL/6N ESC lines (International Knockout Mouse Consortium. ESC germline transmission was observed in 9/11 (82% lines using PH blastocysts, compared to 6/11 (55% when conventional host blastocysts were used. Furthermore, less than 35% (83/240 of mice born in the first litters from conventional chimeras were confirmed to be of ESC-origin. By comparison, 100% (137/137 of the first litter offspring of PH chimeras were confirmed as ESC-derived. Together, these data demonstrate that the PH approach increases the probability of germline transmission and speeds the generation of ESC derived animals from chimeras. Collectively, this approach reduces the time and costs inherent in the

  6. Impacts of CD44 knockdown in cancer cells on tumor and host metabolic systems revealed by quantitative imaging mass spectrometry.

    Science.gov (United States)

    Ohmura, Mitsuyo; Hishiki, Takako; Yamamoto, Takehiro; Nakanishi, Tsuyoshi; Kubo, Akiko; Tsuchihashi, Kenji; Tamada, Mayumi; Toue, Sakino; Kabe, Yasuaki; Saya, Hideyuki; Suematsu, Makoto

    2015-04-30

    CD44 expressed in cancer cells was shown to stabilize cystine transporter (xCT) that uptakes cystine and excretes glutamate to supply cysteine as a substrate for reduced glutathione (GSH) for survival. While targeting CD44 serves as a potentially therapeutic stratagem to attack cancer growth and chemoresistance, the impact of CD44 targeting in cancer cells on metabolic systems of tumors and host tissues in vivo remains to be fully determined. This study aimed to reveal effects of CD44 silencing on alterations in energy metabolism and sulfur-containing metabolites in vitro and in vivo using capillary electrophoresis-mass spectrometry and quantitative imaging mass spectrometry (Q-IMS), respectively. In an experimental model of xenograft transplantation of human colon cancer HCT116 cells in superimmunodeficient NOG mice, snap-frozen liver tissues containing metastatic tumors were examined by Q-IMS. As reported previously, short hairpin CD44 RNA interference (shCD44) in cancer cells caused significant regression of tumor growth in the host liver. Under these circumstances, the CD44 knockdown suppressed polyamines, GSH and energy charges not only in metastatic tumors but also in the host liver. In culture, HCT116 cells treated with shCD44 decreased total amounts of methionine-pool metabolites including spermidine and spermine, and reactive cysteine persulfides, suggesting roles of these metabolites for cancer growth. Collectively, these results suggest that CD44 expressed in cancer accounts for a key regulator of metabolic interplay between tumor and the host tissue. PMID:25461272

  7. Cell alterations induced by a biotherapic for influenza

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    José Nelson Couceiro

    2011-07-01

    Full Text Available Introduction: Influenza viruses have been responsible for highly contagious acute respiratory illnesses with high mortality, mainly in the elderly, which encourages the development of new drugs for the treatment of human flu. The biotherapics are medicines prepared from biological products, which are not chemically defined. They are compounded following the homeopathic procedures indicated for infectious diseases with known etiology [1]. Aim: The purpose of the present study is to verify cellular alterations induced by a biotherapic prepared from the infectious influenza A virus. Methodology: This biotherapic was prepared for this study in the homeopathic potency of 30X according to the Brazilian Homeopathic Pharmacopeia [2]. The concentration of 10% was not cytotoxic to cells, as verified by neutral red assay. The cellular alterations observed in MDCK cells were analyzed by optical microscopy for the quantification of mitosis, nucleoli and lipid bodies. The mitochondrial activity was assessed by MTT assay and the phosphosfructokinase-1 (PFK-1 enzyme activity was analyzed on the MDCK cells treated for 5, 10 and 30 days. Macrophages J778.G8 were treated with this biotherapic to evaluate the immunostimulatory cytokine release. Results: The cellular alterations observed in MDCK cells were verified by optical microscopy. The number of lipid bodies present in MDCK cells stimulated for 10 days was significantly lower (p <0.05 when compared to controls. The biotherapic significantly increased (p <0.05 the number of mitosis and the mitochondrial activity of MDCK cells stimulated for 10 and 30 days. These changes were confirmed by a significant reduction (p <0.05 on the PFK-1 activity. These results suggest that the biotherapic was able to activate the Krebs cycle and pentose-phosphate metabolism to the generation of amino acids and nucleotides, situations common to cells whose rate of mitosis is increased. The quantification of immunostimulatory

  8. Host cell capable of producing enzymes useful for degradation of lignocellulosic material

    Science.gov (United States)

    Los, Alrik Pieter; Sagt, Cornelis Maria Jacobus; Schooneveld-Bergmans, Margot Elisabeth Francoise; Damveld, Robbertus Antonius

    2015-08-18

    The invention relates to a host cell comprising at least four different heterologous polynucleotides chosen from the group of polynucleotides encoding cellulases, hemicellulases and pectinases, wherein the host cell is capable of producing the at least four different enzymes chosen from the group of cellulases, hemicellulases and pectinases, wherein the host cell is a filamentous fungus and is capable of secretion of the at least four different enzymes. This host cell can suitably be used for the production of an enzyme composition that can be used in a process for the saccharification of cellulosic material.

  9. Genetic alterations in head and neck squamous cell carcinomas

    Directory of Open Access Journals (Sweden)

    Nagai M.A.

    1999-01-01

    Full Text Available The genetic alterations observed in head and neck cancer are mainly due to oncogene activation (gain of function mutations and tumor suppressor gene inactivation (loss of function mutations, leading to deregulation of cell proliferation and death. These genetic alterations include gene amplification and overexpression of oncogenes such as myc, erbB-2, EGFR and cyclinD1 and mutations, deletions and hypermethylation leading to p16 and TP53 tumor suppressor gene inactivation. In addition, loss of heterozygosity in several chromosomal regions is frequently observed, suggesting that other tumor suppressor genes not yet identified could be involved in the tumorigenic process of head and neck cancers. The exact temporal sequence of the genetic alterations during head and neck squamous cell carcinoma (HNSCC development and progression has not yet been defined and their diagnostic or prognostic significance is controversial. Advances in the understanding of the molecular basis of head and neck cancer should help in the identification of new markers that could be used for the diagnosis, prognosis and treatment of the disease.

  10. Cutaneous graft-versus-host disease after hematopoietic stem cell transplant - a review*

    Science.gov (United States)

    Villarreal, Cesar Daniel Villarreal; Alanis, Julio Cesar Salas; Pérez, Jose Carlos Jaime; Candiani, Jorge Ocampo

    2016-01-01

    Graft-versus-host disease (GVHD) is a major complication of allogeneic hematopoietic stem cell transplants (allo-HSCT) associated with significant morbidity and mortality. The earliest and most common manifestation is cutaneous graft-versus-host disease. This review focuses on the pathophysiology, clinical features, prevention and treatment of cutaneous graft-versus-host disease. We discuss various insights into the disease's mechanisms and the different treatments for acute and chronic skin graft-versus-host disease. PMID:27438202

  11. Altered B cell receptor signaling in human systemic lupus erythematosus

    Science.gov (United States)

    Jenks, Scott A.; Sanz, Iñaki

    2009-01-01

    Regulation of B cell receptor signaling is essential for the development of specific immunity while retaining tolerance to self. Systemic lupus erythematosus (SLE) is characterized by a loss of B cell tolerance and the production of anti-self antibodies. Accompanying this break down in tolerance are alterations in B cell receptor signal transduction including elevated induced calcium responses and increased protein phosphorylation. Specific pathways that negatively regulate B cell signaling have been shown to be impaired in some SLE patients. These patients have reduced levels of the kinase Lyn in lipid raft microdomains and this reduction is inversely correlated with increased CD45 in lipid rafts. Function and expression of the inhibitory immunoglobulin receptor FcγRIIB is also reduced in Lupus IgM- CD27+ memory cells. Because the relative contribution of different memory and transitional B cell subsets can be abnormal in SLE patients, we believe studies targeted to well defined B cell subsets will be necessary to further our understanding of signaling abnormalities in SLE. Intracellular flow cytometric analysis of signaling is a useful approach to accomplish this goal. PMID:18723129

  12. Alterations induced in Escherichia Coli cells by gamma radiation

    Energy Technology Data Exchange (ETDEWEB)

    Kappke, J.; Schelin, H.R.; Paschuk, S.A.; Denyak, V.; Silva, E.R. da [Federal University of Technology of Parana (CPGEI/UTFPR), Curitiba, PR (Brazil)]. E-mails: jaquekap@yahoo.com.br; schelin@cpgei.cefetpr.br; sergei@utfpr.edu.br; Jesus, E.F.O. de; Lopes, R.T. [Universidade Federal do Rio de Janeiro (UFRJ), RJ (Brazil). Coordenacao dos Programas de Pos-graduacao de Engenharia (COPPE). Lab. de Instrumentacao Nuclear]. E-mails: ricardo@lin.ufrj.br; edgar@lin.ufrj.br; Carlin, N.; Toledo, E.S. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Fisica]. E-mail: nelson.carlin@dfn.if.usp.br

    2007-07-01

    Modifications occurred in Escherichia coli cells exposed to gamma radiation ({sup 60}Co source) were investigated. The irradiations were done at the LIN-COPPE laboratory of the UFRJ and the analysis at the Biology Department of the UTFPR. The E. coli cells were irradiated with 30, 60, 90, 120, 150, 180, 210, 240, 300, 480, 600 e 750 Gy doses. The samples were analyzed with Gram-stain, biochemical tests in EPM, MIO and Lysine Broth, Simmons Cytrate Medium and Rhamnose Broth, antibiogram and isolation of auxotrophic mutants. It was observed that for the received doses the E. coli did not show morphological alterations in the tests. Some E. Coli cells showed to be able to deaminade the L-tryptophan or they changed their sensibility for amoxillin and cephaloonine after the irradiation. The existence of aauxotrophic mutants after irradiation was also verified. (author)

  13. Host Cell Autophagy Modulates Early Stages of Adenovirus Infections in Airway Epithelial Cells

    OpenAIRE

    Zeng, Xuehuo; Carlin, Cathleen R

    2013-01-01

    Human adenoviruses typically cause mild infections in the upper or lower respiratory tract, gastrointestinal tract, or ocular epithelium. However, adenoviruses may be life-threatening in patients with impaired immunity and some serotypes cause epidemic outbreaks. Attachment to host cell receptors activates cell signaling and virus uptake by endocytosis. At present, it is unclear how vital cellular homeostatic mechanisms affect these early steps in the adenovirus life cycle. Autophagy is a lys...

  14. Identification and monitoring of host cell proteins by mass spectrometry combined with high performance immunochemistry testing.

    Directory of Open Access Journals (Sweden)

    Katrin Bomans

    Full Text Available Biotherapeutics are often produced in non-human host cells like Escherichia coli, yeast, and various mammalian cell lines. A major focus of any therapeutic protein purification process is to reduce host cell proteins to an acceptable low level. In this study, various E. coli host cell proteins were identified at different purifications steps by HPLC fractionation, SDS-PAGE analysis, and tryptic peptide mapping combined with online liquid chromatography mass spectrometry (LC-MS. However, no host cell proteins could be verified by direct LC-MS analysis of final drug substance material. In contrast, the application of affinity enrichment chromatography prior to comprehensive LC-MS was adequate to identify several low abundant host cell proteins at the final drug substance level. Bacterial alkaline phosphatase (BAP was identified as being the most abundant host cell protein at several purification steps. Thus, we firstly established two different assays for enzymatic and immunological BAP monitoring using the cobas® technology. By using this strategy we were able to demonstrate an almost complete removal of BAP enzymatic activity by the established therapeutic protein purification process. In summary, the impact of fermentation, purification, and formulation conditions on host cell protein removal and biological activity can be conducted by monitoring process-specific host cell proteins in a GMP-compatible and high-throughput (> 1000 samples/day manner.

  15. Severe insulin resistance alters metabolism in mesenchymal progenitor cells.

    Science.gov (United States)

    Balhara, Bharti; Burkart, Alison; Topcu, Vehap; Lee, Youn-Kyoung; Cowan, Chad; Kahn, C Ronald; Patti, Mary-Elizabeth

    2015-06-01

    Donohue syndrome (DS) is characterized by severe insulin resistance due to mutations in the insulin receptor (INSR) gene. To identify molecular defects contributing to metabolic dysregulation in DS in the undifferentiated state, we generated mesenchymal progenitor cells (MPCs) from induced pluripotent stem cells derived from a 4-week-old female with DS and a healthy newborn male (control). INSR mRNA and protein were significantly reduced in DS MPC (for β-subunit, 64% and 89% reduction, respectively, P consumption in both the basal state (87% higher, P =.09) and in response to the uncoupler carbonyl cyanide-p-triflouromethoxyphenylhydrazone (2-fold increase, P =.06). Although mitochondrial DNA or mass did not differ, oxidative phosphorylation protein complexes III and V were increased in DS (by 37% and 6%, respectively; P < .05). Extracellular acidification also tended to increase in DS (91% increase, P = .07), with parallel significant increases in lactate secretion (34% higher at 4 h, P < .05). In summary, DS MPC maintain signaling downstream of the INSR, suggesting that IGF-1R signaling may partly compensate for INSR mutations. However, alterations in receptor expression and pathway-specific defects in insulin signaling, even in undifferentiated cells, can alter cellular oxidative metabolism, potentially via transcriptional mechanisms. PMID:25811318

  16. Diabetes alters intracellular calcium transients in cardiac endothelial cells.

    Directory of Open Access Journals (Sweden)

    Abdul Q Sheikh

    Full Text Available Diabetic cardiomyopathy (DCM is a diabetic complication, which results in myocardial dysfunction independent of other etiological factors. Abnormal intracellular calcium ([Ca(2+](i homeostasis has been implicated in DCM and may precede clinical manifestation. Studies in cardiomyocytes have shown that diabetes results in impaired [Ca(2+](i homeostasis due to altered sarcoplasmic reticulum Ca(2+ ATPase (SERCA and sodium-calcium exchanger (NCX activity. Importantly, altered calcium homeostasis may also be involved in diabetes-associated endothelial dysfunction, including impaired endothelium-dependent relaxation and a diminished capacity to generate nitric oxide (NO, elevated cell adhesion molecules, and decreased angiogenic growth factors. However, the effect of diabetes on Ca(2+ regulatory mechanisms in cardiac endothelial cells (CECs remains unknown. The objective of this study was to determine the effect of diabetes on [Ca(2+](i homeostasis in CECs in the rat model (streptozotocin-induced of DCM. DCM-associated cardiac fibrosis was confirmed using picrosirius red staining of the myocardium. CECs isolated from the myocardium of diabetic and wild-type rats were loaded with Fura-2, and UTP-evoked [Ca(2+](i transients were compared under various combinations of SERCA, sarcoplasmic reticulum Ca(2+ ATPase (PMCA and NCX inhibitors. Diabetes resulted in significant alterations in SERCA and NCX activities in CECs during [Ca(2+](i sequestration and efflux, respectively, while no difference in PMCA activity between diabetic and wild-type cells was observed. These results improve our understanding of how diabetes affects calcium regulation in CECs, and may contribute to the development of new therapies for DCM treatment.

  17. Patterns of oligonucleotide sequences in viral and host cell RNA identify mediators of the host innate immune system.

    Directory of Open Access Journals (Sweden)

    Benjamin D Greenbaum

    Full Text Available The innate immune response provides a first line of defense against pathogens by targeting generic differential features that are present in foreign organisms but not in the host. These innate responses generate selection forces acting both in pathogens and hosts that further determine their co-evolution. Here we analyze the nucleic acid sequence fingerprints of these selection forces acting in parallel on both host innate immune genes and ssRNA viral genomes. We do this by identifying dinucleotide biases in the coding regions of innate immune response genes in plasmacytoid dendritic cells, and then use this signal to identify other significant host innate immune genes. The persistence of these biases in the orthologous groups of genes in humans and chickens is also examined. We then compare the significant motifs in highly expressed genes of the innate immune system to those in ssRNA viruses and study the evolution of these motifs in the H1N1 influenza genome. We argue that the significant under-represented motif pattern of CpG in an AU context--which is found in both the ssRNA viruses and innate genes, and has decreased throughout the history of H1N1 influenza replication in humans--is immunostimulatory and has been selected against during the co-evolution of viruses and host innate immune genes. This shows how differences in host immune biology can drive the evolution of viruses that jump into species with different immune priorities than the original host.

  18. Impact of host cell line adaptation on quasispecies composition and glycosylation of influenza A virus hemagglutinin.

    Directory of Open Access Journals (Sweden)

    Jana Verena Roedig

    Full Text Available The genome of influenza A viruses is constantly changing (genetic drift resulting in small, gradual changes in viral proteins. Alterations within antibody recognition sites of the viral membrane glycoproteins hemagglutinin (HA and neuraminidase (NA result in an antigenetic drift, which requires the seasonal update of human influenza virus vaccines. Generally, virus adaptation is necessary to obtain sufficiently high virus yields in cell culture-derived vaccine manufacturing. In this study detailed HA N-glycosylation pattern analysis was combined with in-depth pyrosequencing analysis of the virus genomic RNA. Forward and backward adaptation from Madin-Darby Canine Kidney (MDCK cells to African green monkey kidney (Vero cells was investigated for two closely related influenza A virus PR/8/34 (H1N1 strains: from the National Institute for Biological Standards and Control (NIBSC or the Robert Koch Institute (RKI. Furthermore, stability of HA N-glycosylation patterns over ten consecutive passages and different harvest time points is demonstrated. Adaptation to Vero cells finally allowed efficient influenza A virus replication in Vero cells. In contrast, during back-adaptation the virus replicated well from the very beginning. HA N-glycosylation patterns were cell line dependent and stabilized fast within one (NIBSC-derived virus or two (RKI-derived virus successive passages during adaptation processes. However, during adaptation new virus variants were detected. These variants carried "rescue" mutations on the genomic level within the HA stem region, which result in amino acid substitutions. These substitutions finally allowed sufficient virus replication in the new host system. According to adaptation pressure the composition of the virus populations varied. In Vero cells a selection for "rescue" variants was characteristic. After back-adaptation to MDCK cells some variants persisted at indifferent frequencies, others slowly diminished and even

  19. Salinity regulation of the interaction of halovirus SNJ1 with its host and alteration of the halovirus replication strategy to adapt to the variable ecosystem.

    Directory of Open Access Journals (Sweden)

    Yunjun Mei

    Full Text Available Halovirus is a major force that affects the evolution of extreme halophiles and the biogeochemistry of hypersaline environments. However, until now, the systematic studies on the halovirus ecology and the effects of salt concentration on virus-host systems are lacking. To provide more valuable information for understanding ecological strategies of a virus-host system in the hypersaline ecosystem, we studied the interaction between halovirus SNJ1 and its host Natrinema sp.J7-2 under various NaCl concentrations. We found that the adsorption rate and lytic rate increased with salt concentration, demonstrating that a higher salt concentration promoted viral adsorption and proliferation. Contrary to the lytic rate, the lysogenic rate decreased as the salt concentration increased. Our results also demonstrated that cells incubated at a high salt concentration prior to infection increased the ability of the virus to adsorb and lyse its host cells; therefore, the physiological status of host cells also affected the virus-host interaction. In conclusion, SNJ1 acted as a predator, lysing host cells and releasing progeny viruses in hypersaline environments; in low salt environments, viruses lysogenized host cells to escape the damage from low salinity.

  20. Salinity regulation of the interaction of halovirus SNJ1 with its host and alteration of the halovirus replication strategy to adapt to the variable ecosystem.

    Science.gov (United States)

    Mei, Yunjun; He, Congcong; Huang, Yongchi; Liu, Ying; Zhang, Ziqian; Chen, Xiangdong; Shen, Ping

    2015-01-01

    Halovirus is a major force that affects the evolution of extreme halophiles and the biogeochemistry of hypersaline environments. However, until now, the systematic studies on the halovirus ecology and the effects of salt concentration on virus-host systems are lacking. To provide more valuable information for understanding ecological strategies of a virus-host system in the hypersaline ecosystem, we studied the interaction between halovirus SNJ1 and its host Natrinema sp.J7-2 under various NaCl concentrations. We found that the adsorption rate and lytic rate increased with salt concentration, demonstrating that a higher salt concentration promoted viral adsorption and proliferation. Contrary to the lytic rate, the lysogenic rate decreased as the salt concentration increased. Our results also demonstrated that cells incubated at a high salt concentration prior to infection increased the ability of the virus to adsorb and lyse its host cells; therefore, the physiological status of host cells also affected the virus-host interaction. In conclusion, SNJ1 acted as a predator, lysing host cells and releasing progeny viruses in hypersaline environments; in low salt environments, viruses lysogenized host cells to escape the damage from low salinity. PMID:25853566

  1. Proteomics analysis of EV71-infected cells reveals the involvement of host protein NEDD4L in EV71 replication.

    Science.gov (United States)

    Kuo, Rei-Lin; Lin, Ya-Han; Wang, Robert Yung-Liang; Hsu, Chia-Wei; Chiu, Yi-Ting; Huang, Hsing-I; Kao, Li-Ting; Yu, Jau-Song; Shih, Shin-Ru; Wu, Chih-Ching

    2015-04-01

    Enterovirus 71 (EV71) is a human enterovirus that has seriously affected the Asia-Pacific area for the past two decades. EV71 infection can result in mild hand-foot-and-mouth disease and herpangina and may occasionally lead to severe neurological complications in children. However, the specific biological processes that become altered during EV71 infection remain unclear. To further explore host responses upon EV71 infection, we identified proteins differentially expressed in EV71-infected human glioblastoma SF268 cells using isobaric mass tag (iTRAQ) labeling coupled with multidimensional liquid chromatography-mass spectrometry (LC-MS/MS). Network analysis of proteins altered in cells infected with EV71 revealed that the changed biological processes are related to protein and ion transport, regulation of protein degradation, and homeostatic processes. We confirmed that the levels of NEDD4L and PSMF1 were increased and reduced, respectively, in EV71-infected cells compared to mock-infected control cells. To determine the physiological relevance of our findings, we investigated the consequences of EV71 infection in cells with NEDD4L or PSMF1 depletion. We found that the depletion of NEDD4L significantly reduced the replication of EV71, whereas PSMF1 knockdown enhanced EV71 replication. Collectively, our findings provide the first evidence of proteome-wide dysregulation by EV71 infection and suggest a novel role for the host protein NEDD4L in the replication of this virus.

  2. Host Factors Invovled in the Entry of Coronaviruses into Mammalian Cells

    NARCIS (Netherlands)

    Burkard, C.

    2015-01-01

    Enveloped viruses need to fuse with a host cell membrane in order to deliver their genome into the host cell. While some viruses fuse with the plasma membrane, many viruses are endocytosed prior to fusion. Specific cues in the endosomal microenvironment induce conformational changes in the viral fus

  3. Cyanobacterium sp. host cell and vector for production of chemical compounds in Cyanobacterial cultures

    Energy Technology Data Exchange (ETDEWEB)

    Piven, Irina; Friedrich, Alexandra; Duhring, Ulf; Uliczka, Frank; Baier, Kerstin; Inaba, Masami; Shi, Tuo; Wang, Kui; Enke, Heike; Kramer, Dan

    2016-04-19

    A cyanobacterial host cell, Cyanobacterium sp., that harbors at least one recombinant gene for the production of a chemical compounds is provided, as well as vectors derived from an endogenous plasmid isolated from the cell.

  4. Cyanobacterium sp. host cell and vector for production of chemical compounds in cyanobacterial cultures

    Science.gov (United States)

    Piven, Irina; Friedrich, Alexandra; Duhring, Ulf; Uliczka, Frank; Baier, Kerstin; Inaba, Masami; Shi, Tuo; Wang, Kui; Enke, Heike; Kramer, Dan

    2014-09-30

    A cyanobacterial host cell, Cyanobacterium sp., that harbors at least one recombinant gene for the production of a chemical compounds is provided, as well as vectors derived from an endogenous plasmid isolated from the cell.

  5. From microbiology to cell biology: when an intracellular bacterium becomes part of its host cell.

    Science.gov (United States)

    McCutcheon, John P

    2016-08-01

    Mitochondria and chloroplasts are now called organelles, but they used to be bacteria. As they transitioned from endosymbionts to organelles, they became more and more integrated into the biochemistry and cell biology of their hosts. Work over the last 15 years has shown that other symbioses show striking similarities to mitochondria and chloroplasts. In particular, many sap-feeding insects house intracellular bacteria that have genomes that overlap mitochondria and chloroplasts in terms of size and coding capacity. The massive levels of gene loss in some of these bacteria suggest that they, too, are becoming highly integrated with their host cells. Understanding these bacteria will require inspiration from eukaryotic cell biology, because a traditional microbiological framework is insufficient for understanding how they work.

  6. Genetic alterations in B-cell non-Hodgkin's lymphoma

    Directory of Open Access Journals (Sweden)

    Magić Zvonko

    2005-01-01

    Full Text Available Background. Although the patients with diagnosed B-NHL are classified into the same disease stage on the basis of clinical, histopathological, and immunological parameters, they respond significantly different to the applied treatment. This points out the possibility that within the same group of lymphoma there are different diseases at molecular level. For that reason many studies deal with the detection of gene alterations in lymphomas to provide a better framework for diagnosis and treatment of these hematological malignancies. Aim. To define genetic alterations in the B-NHL with highest possibilities for diagnostic purposes and molecular detection of MRD. Methods. Formalin fixed and paraffin embedded lymph node tissues from 45 patients were examined by different PCR techniques for the presence of IgH and TCR γ gene rearrangement; K-ras and H-ras mutations; c-myc amplification and bcl-2 translocation. There were 34 cases of B-cell non-Hodgkin’s lymphoma (B-NHL, 5 cases of T-cell non-Hodgkin’s lymphoma (T-NHL and 6 cases of chronic lymphadenitis (CL. The mononuclear cell fraction of the peripheral blood of 12 patients with B-NHL was analyzed for the presence of monoclonality at the time of diagnosis and in 3 to 6 months time intervals after an autologous bone marrow transplantation (BMT. Results. The monoclonality of B-lymphocytes, as evidenced by DNA fragment length homogeneity, was detected in 88 % (30/34 of B-NHL, but never in CL, T-NHL, or in normal PBL. Bcl-2 translocation was detected in 7/31 (22.6% B-NHL specimens, c-myc amplification 9/31 (29%, all were more than doubled, K-ras mutations in 1/31 (3.23% and H-ras mutations in 2/31 (6.45% of the examined B-NHL samples. In the case of LC and normal PBL, however, these gene alterations were not detected. All the patients (12 with B-NHL had dominant clone of B-lymphocyte in the peripheral blood at the time of diagnosis while only in 2 of 12 patients MRD was detected 3 or 6 months after

  7. Graphene-Based Interfaces Do Not Alter Target Nerve Cells.

    Science.gov (United States)

    Fabbro, Alessandra; Scaini, Denis; León, Verónica; Vázquez, Ester; Cellot, Giada; Privitera, Giulia; Lombardi, Lucia; Torrisi, Felice; Tomarchio, Flavia; Bonaccorso, Francesco; Bosi, Susanna; Ferrari, Andrea C; Ballerini, Laura; Prato, Maurizio

    2016-01-26

    Neural-interfaces rely on the ability of electrodes to transduce stimuli into electrical patterns delivered to the brain. In addition to sensitivity to the stimuli, stability in the operating conditions and efficient charge transfer to neurons, the electrodes should not alter the physiological properties of the target tissue. Graphene is emerging as a promising material for neuro-interfacing applications, given its outstanding physico-chemical properties. Here, we use graphene-based substrates (GBSs) to interface neuronal growth. We test our GBSs on brain cell cultures by measuring functional and synaptic integrity of the emerging neuronal networks. We show that GBSs are permissive interfaces, even when uncoated by cell adhesion layers, retaining unaltered neuronal signaling properties, thus being suitable for carbon-based neural prosthetic devices. PMID:26700626

  8. Simulated Hypergravity Alters Vascular Smooth Muscle Cell Proliferation and Motility

    Science.gov (United States)

    Hunt, Shameka; Bettis, Barika; Harris-Hooker, Sandra; Sanford, Gary L.

    1997-01-01

    The cellular effects of gravity are poorly understood due to its constancy and nonavailability of altered gravitational models. Such an understanding is crucial for prolonged space flights. In these studies, we assessed the influence of centrifugation at 6G (HGrav) on vascular smooth muscle (SMC) mobility and proliferation. Cells were: (a) plated at low density and subjected to HGrav for 24-72 hr for proliferation studies, or (b) grown to confluency, subjected to HGrav, mechanically denuded and monitored for cell movement into the denuded area. Controls were maintained under normogravity. SMC showed a 50% inhibition of growth under HGrav and 10% serum; HGrav and low serum resulted in greater growth inhibition. The rate of movement of SMC into the denuded area was 2-3-fold higher under HGrav in low serum compared to controls, but similar in 10% serum. These studies show that HGrav has significant effects on SMC growth and mobility, which are dependent on serum levels.

  9. Hypertension alters phosphorylation of VASP in brain endothelial cells.

    Science.gov (United States)

    Arlier, Zulfikar; Basar, Murat; Kocamaz, Erdogan; Kiraz, Kemal; Tanriover, Gamze; Kocer, Gunnur; Arlier, Sefa; Giray, Semih; Nasırcılar, Seher; Gunduz, Filiz; Senturk, Umit K; Demir, Necdet

    2015-04-01

    Hypertension impairs cerebral vascular function. Vasodilator-stimulated phosphoprotein (VASP) mediates active reorganization of the cytoskeleton via membrane ruffling, aggregation and tethering of actin filaments. VASP regulation of endothelial barrier function has been demonstrated by studies using VASP(-/-) animals under conditions associated with tissue hypoxia. We hypothesize that hypertension regulates VASP expression and/or phosphorylation in endothelial cells, thereby contributing to dysfunction in the cerebral vasculature. Because exercise has direct and indirect salutary effects on vascular systems that have been damaged by hypertension, we also investigated the effect of exercise on maintenance of VASP expression and/or phosphorylation. We used immunohistochemistry, Western blotting and immunocytochemistry to examine the effect of hypertension on VASP expression and phosphorylation in brain endothelial cells in normotensive [Wistar-Kyoto (WKY)] and spontaneously hypertensive (SH) rats under normal and exercise conditions. In addition, we analyzed VASP regulation in normoxia- and hypoxia-induced endothelial cells. Brain endothelial cells exhibited significantly lower VASP immunoreactivity and phosphorylation at the Ser157 residue in SHR versus WKY rats. Exercise reversed hypertension-induced alterations in VASP phosphorylation. Western blotting and immunocytochemistry indicated reduction in VASP phosphorylation in hypoxic versus normoxic endothelial cells. These results suggest that diminished VASP expression and/or Ser157 phosphorylation mediates endothelial changes associated with hypertension and exercise may normalize these changes, at least in part, by restoring VASP phosphorylation. PMID:24894047

  10. Gestational protein restriction alters cell proliferation in rat placenta.

    Science.gov (United States)

    Rebelato, Hércules Jonas; Esquisatto, Marcelo Augusto Marreto; de Sousa Righi, Eloá Fernanda; Catisti, Rosana

    2016-04-01

    We recently showed that gestational protein restriction (GPR) alters the structure of the rat placenta on day 19 of gestation (dG). The aim of the study was to investigate the spatial and temporal immunolocalization of proliferating cell antigen Ki67 in normal and GPR placental development. Pregnant Wistar rats were divided into two groups: normal (NP, 17 % casein) or low-protein diet (LP, 6 % casein). Placentas and fetus were collected and weighed at 15, 17, 19 and 21 dG. Morphological, morphometric and ultrastructural analyses were performed. Immunoperoxidase was used to identify nuclear antigen Ki67 in placental sections. We observed a significant reduction in the number of trophoblast giant cells and glycogen cells in the LP group. Placental weight was significantly reduced only at 17 dG in the LP group, in parallel to a decrease in glycogen cells. From 15 to 21 dG, the thickness of the junctional zone (JZ) decreased in NP and LP animals, while that of the labyrinth zone (LZ) increased in parallel to a reduction in the number of proliferating cells in this LZ zone. GPR significantly inhibits cell proliferation in the JZ, especially at 15 and 17 dG. The ultrastructural appearance of the cytoplasm of giant and cytotrophoblastic cells indicates degeneration from 15 to 21 dG and this effect is enhanced in LP animals suggesting early aging. Offspring of NP dams were significantly heavier than offspring of LP dams at 21 dG. GPR causes modifications in specific regions of the placenta, cell proliferation inhibition and fetal growth restriction.

  11. Alterations of proteins in MDCK cells during acute potassium deficiency.

    Science.gov (United States)

    Peerapen, Paleerath; Ausakunpipat, Nardtaya; Chanchaem, Prangwalai; Thongboonkerd, Visith

    2016-06-01

    Chronic K(+) deficiency can cause hypokalemic nephropathy associated with metabolic alkalosis, polyuria, tubular dilatation, and tubulointerstitial injury. However, effects of acute K(+) deficiency on the kidney remained unclear. This study aimed to explore such effects by evaluating changes in levels of proteins in renal tubular cells during acute K(+) deficiency. MDCK cells were cultivated in normal K(+) (NK) (K(+)=5.3 mM), low K(+) (LK) (K(+)=2.5 mM), or K(+) depleted (KD) (K(+)=0 mM) medium for 24 h and then harvested. Cellular proteins were resolved by two-dimensional gel electrophoresis (2-DE) and visualized by SYPRO Ruby staining (5 gels per group). Spot matching and quantitative intensity analysis revealed a total 48 protein spots that had significantly differential levels among the three groups. Among these, 46 and 30 protein spots had differential levels in KD group compared to NK and LK groups, respectively. Comparison between LK and NK groups revealed only 10 protein spots that were differentially expressed. All of these differentially expressed proteins were successfully identified by Q-TOF MS and/or MS/MS analyses. The altered levels of heat shock protein 90 (HSP90), ezrin, lamin A/C, tubulin, chaperonin-containing TCP1 (CCT1), and calpain 1 were confirmed by Western blot analysis. Global protein network analysis showed three main functional networks, including 1) cell growth and proliferation, 2) cell morphology, cellular assembly and organization, and 3) protein folding in which the altered proteins were involved. Further investigations on these networks may lead to better understanding of pathogenic mechanisms of low K(+)-induced renal injury.

  12. Cell-to-cell communication and cellular environment alter the somatostatin status of delta cells

    International Nuclear Information System (INIS)

    Research highlights: → TGP52 cells display enhanced functionality in pseudoislet form. → Somatostatin content was reduced, but secretion increased in high glucose conditions. → Cellular interactions and environment alter the somatostatin status of TGP52 cells. -- Abstract: Introduction: Somatostatin, released from pancreatic delta cells, is a potent paracrine inhibitor of insulin and glucagon secretion. Islet cellular interactions and glucose homeostasis are essential to maintain normal patterns of insulin secretion. However, the importance of cell-to-cell communication and cellular environment in the regulation of somatostatin release remains unclear. Methods: This study employed the somatostatin-secreting TGP52 cell line maintained in DMEM:F12 (17.5 mM glucose) or DMEM (25 mM glucose) culture media. The effect of pseudoislet formation and culture medium on somatostatin content and release in response to a variety of stimuli was measured by somatostatin EIA. In addition, the effect of pseudoislet formation on cellular viability (MTT and LDH assays) and proliferation (BrdU ELISA) was determined. Results: TGP52 cells readily formed pseudoislets and showed enhanced functionality in three-dimensional form with increased E-cadherin expression irrespective of the culture environment used. However, culture in DMEM decreased cellular somatostatin content (P < 0.01) and increased somatostatin secretion in response to a variety of stimuli including arginine, calcium and PMA (P < 0.001) when compared with cells grown in DMEM:F12. Configuration of TGP52 cells as pseudoislets reduced the proliferative rate and increased cellular cytotoxicity irrespective of culture medium used. Conclusions: Somatostatin secretion is greatly facilitated by cell-to-cell interactions and E-cadherin expression. Cellular environment and extracellular glucose also significantly influence the function of delta cells.

  13. Phenotypic and Functional Alterations in Circulating Memory CD8 T Cells with Time after Primary Infection.

    Directory of Open Access Journals (Sweden)

    Matthew D Martin

    2015-10-01

    Full Text Available Memory CD8 T cells confer increased protection to immune hosts upon secondary viral, bacterial, and parasitic infections. The level of protection provided depends on the numbers, quality (functional ability, and location of memory CD8 T cells present at the time of infection. While primary memory CD8 T cells can be maintained for the life of the host, the full extent of phenotypic and functional changes that occur over time after initial antigen encounter remains poorly characterized. Here we show that critical properties of circulating primary memory CD8 T cells, including location, phenotype, cytokine production, maintenance, secondary proliferation, secondary memory generation potential, and mitochondrial function change with time after infection. Interestingly, phenotypic and functional alterations in the memory population are not due solely to shifts in the ratio of effector (CD62Llo and central memory (CD62Lhi cells, but also occur within defined CD62Lhi memory CD8 T cell subsets. CD62Lhi memory cells retain the ability to efficiently produce cytokines with time after infection. However, while it is was not formally tested whether changes in CD62Lhi memory CD8 T cells over time occur in a cell intrinsic manner or are due to selective death and/or survival, the gene expression profiles of CD62Lhi memory CD8 T cells change, phenotypic heterogeneity decreases, and mitochondrial function and proliferative capacity in either a lymphopenic environment or in response to antigen re-encounter increase with time. Importantly, and in accordance with their enhanced proliferative and metabolic capabilities, protection provided against chronic LCMV clone-13 infection increases over time for both circulating memory CD8 T cell populations and for CD62Lhi memory cells. Taken together, the data in this study reveal that memory CD8 T cells continue to change with time after infection and suggest that the outcome of vaccination strategies designed to elicit

  14. HHV-8 encoded LANA-1 alters the higher organization of the cell nucleus

    Directory of Open Access Journals (Sweden)

    Klein George

    2007-04-01

    Full Text Available Abstract The latency-associated nuclear antigen (LANA-1 of Human Herpes Virus 8 (HHV-8, alternatively called Kaposi Sarcoma Herpes Virus (KSHV is constitutively expressed in all HHV-8 infected cells. LANA-1 accumulates in well-defined foci that co-localize with the viral episomes. We have previously shown that these foci are tightly associated with the borders of heterochromatin 1. We have also shown that exogenously expressed LANA-1 causes an extensive re-organization of Hoechst 33248 DNA staining patterns of the nuclei in non-HHV-8 infected cells 2. Here we show that this effect includes the release of the bulk of DNA from heterochromatic areas, in both human and mouse cells, without affecting the overall levels of heterochromatin associated histone H3 lysine 9 tri-methylation (3MK9H3. The release of DNA from the heterochromatic chromocenters in LANA-1 transfected mouse cells co-incides with the dispersion of the chromocenter associated methylcytosin binding protein 2 (MECP2. The localization of 3MK9H3 to the remnants of the chromocenters remains unaltered. Moreover, exogeneously expressed LANA-1 leads to the relocation of the chromocenters to the nuclear periphery, indicating extensive changes in the positioning of the chromosomal domains in the LANA-1 harboring interphase nucleus. Using a series of deletion mutants we have shown that the chromatin rearranging effects of LANA-1 require the presence of a short (57 amino acid region that is located immediately upstream of the internal acidic repeats. This sequence lies within the previously mapped binding site to histone methyltransferase SUV39H1. We suggest that the highly concentrated LANA-1, anchored to the host genome in the nuclear foci of latently infected cells and replicated through each cell generation, may function as "epigenetic modifier". The induction of histone modification in adjacent host genes may lead to altered gene expression, thereby contributing to the viral oncogenesis.

  15. Epigenetic alterations differ in phenotypically distinct human neuroblastoma cell lines

    International Nuclear Information System (INIS)

    Epigenetic aberrations and a CpG island methylator phenotype have been shown to be associated with poor outcomes in children with neuroblastoma (NB). Seven cancer related genes (THBS-1, CASP8, HIN-1, TIG-1, BLU, SPARC, and HIC-1) that have been shown to have epigenetic changes in adult cancers and play important roles in the regulation of angiogenesis, tumor growth, and apoptosis were analyzed to investigate the role epigenetic alterations play in determining NB phenotype. Two NB cell lines (tumorigenic LA1-55n and non-tumorigenic LA1-5s) that differ in their ability to form colonies in soft agar and tumors in nude mice were used. Quantitative RNA expression analyses were performed on seven genes in LA1-5s, LA1-55n and 5-Aza-dC treated LA1-55n NB cell lines. The methylation status around THBS-1, HIN-1, TIG-1 and CASP8 promoters was examined using methylation specific PCR. Chromatin immunoprecipitation assay was used to examine histone modifications along the THBS-1 promoter. Luciferase assay was used to determine THBS-1 promoter activity. Cell proliferation assay was used to examine the effect of 5-Aza-dC on NB cell growth. The soft agar assay was used to determine the tumorigenicity. Promoter methylation values for THBS-1, HIN-1, TIG-1, and CASP8 were higher in LA1-55n cells compared to LA1-5s cells. Consistent with the promoter methylation status, lower levels of gene expression were detected in the LA1-55n cells. Histone marks associated with repressive chromatin states (H3K9Me3, H3K27Me3, and H3K4Me3) were identified in the THBS-1 promoter region in the LA1-55n cells, but not the LA1-5s cells. In contrast, the three histone codes associated with an active chromatin state (acetyl H3, acetyl H4, and H3K4Me3) were present in the THBS-1 promoter region in LA1-5s cells, but not the LA1-55n cells, suggesting that an accessible chromatin structure is important for THBS-1 expression. We also show that 5-Aza-dC treatment of LA1-55n cells alters the DNA methylation

  16. Interactions between Trypanosoma cruzi Secreted Proteins and Host Cell Signaling Pathways

    Science.gov (United States)

    Watanabe Costa, Renata; da Silveira, Jose F.; Bahia, Diana

    2016-01-01

    Chagas disease is one of the prevalent neglected tropical diseases, affecting at least 6–7 million individuals in Latin America. It is caused by the protozoan parasite Trypanosoma cruzi, which is transmitted to vertebrate hosts by blood-sucking insects. After infection, the parasite invades and multiplies in the myocardium, leading to acute myocarditis that kills around 5% of untreated individuals. T. cruzi secretes proteins that manipulate multiple host cell signaling pathways to promote host cell invasion. The primary secreted lysosomal peptidase in T. cruzi is cruzipain, which has been shown to modulate the host immune response. Cruzipain hinders macrophage activation during the early stages of infection by interrupting the NF-kB P65 mediated signaling pathway. This allows the parasite to survive and replicate, and may contribute to the spread of infection in acute Chagas disease. Another secreted protein P21, which is expressed in all of the developmental stages of T. cruzi, has been shown to modulate host phagocytosis signaling pathways. The parasite also secretes soluble factors that exert effects on host extracellular matrix, such as proteolytic degradation of collagens. Finally, secreted phospholipase A from T. cruzi contributes to lipid modifications on host cells and concomitantly activates the PKC signaling pathway. Here, we present a brief review of the interaction between secreted proteins from T. cruzi and the host cells, emphasizing the manipulation of host signaling pathways during invasion. PMID:27065960

  17. IMMUNE INHIBITION OF VIRUS RELEASE FROM HUMAN AND NONHUMAN CELLS BY ANTIBODY TO VIRAL AND HOST-CELL DETERMINANTS

    NARCIS (Netherlands)

    SHARIFF, DM; DESPERBASQUES, M; BILLSTROM, M; GEERLIGS, HJ; WELLING, GW; WELLINGWESTER, S; BUCHAN, A; SKINNER, GRB

    1991-01-01

    Immune inhibition of release of the DNA virues, herpes simplex virus types 1 and 2 and pseudorabies virus by anti-viral and anti-host cell sera occurred while two RNA viruses, influenza and encephalomyocarditis, were inhibited only by anti-viral sera (not anti-host cell sera). Simian virus 40 and su

  18. Computational prediction of Mycoplasma hominis proteins targeting in nucleus of host cell and their implication in prostate cancer etiology.

    Science.gov (United States)

    Khan, Shahanavaj; Zakariah, Mohammed; Palaniappan, Sellappan

    2016-08-01

    Cancer has long been assumed to be a genetic disease. However, recent evidence supports the enigmatic connection of bacterial infection with the growth and development of various types of cancers. The cause and mechanism of the growth and development of prostate cancer due to Mycoplasma hominis remain unclear. Prostate cancer cells are infected and colonized by enteroinvasive M. hominis, which controls several factors that can affect prostate cancer growth in susceptible persons. We investigated M. hominis proteins targeting the nucleus of host cells and their implications in prostate cancer etiology. Many vital processes are controlled in the nucleus, where the proteins targeting M. hominis may have various potential implications. A total of 29/563 M. hominis proteins were predicted to target the nucleus of host cells. These include numerous proteins with the capability to alter normal growth activities. In conclusion, our results emphasize that various proteins of M. hominis targeted the nucleus of host cells and were involved in prostate cancer etiology through different mechanisms and strategies.

  19. Alterations of red cell membrane properties in neuroacanthocytosis.

    Directory of Open Access Journals (Sweden)

    Claudia Siegl

    Full Text Available Neuroacanthocytosis (NA refers to a group of heterogenous, rare genetic disorders, namely chorea acanthocytosis (ChAc, McLeod syndrome (MLS, Huntington's disease-like 2 (HDL2 and pantothenate kinase associated neurodegeneration (PKAN, that mainly affect the basal ganglia and are associated with similar neurological symptoms. PKAN is also assigned to a group of rare neurodegenerative diseases, known as NBIA (neurodegeneration with brain iron accumulation, associated with iron accumulation in the basal ganglia and progressive movement disorder. Acanthocytosis, the occurrence of misshaped erythrocytes with thorny protrusions, is frequently observed in ChAc and MLS patients but less prevalent in PKAN (about 10% and HDL2 patients. The pathological factors that lead to the formation of the acanthocytic red blood cell shape are currently unknown. The aim of this study was to determine whether NA/NBIA acanthocytes differ in their functionality from normal erythrocytes. Several flow-cytometry-based assays were applied to test the physiological responses of the plasma membrane, namely drug-induced endocytosis, phosphatidylserine exposure and calcium uptake upon treatment with lysophosphatidic acid. ChAc red cell samples clearly showed a reduced response in drug-induced endovesiculation, lysophosphatidic acid-induced phosphatidylserine exposure, and calcium uptake. Impaired responses were also observed in acanthocyte-positive NBIA (PKAN red cells but not in patient cells without shape abnormalities. These data suggest an "acanthocytic state" of the red cell where alterations in functional and interdependent membrane properties arise together with an acanthocytic cell shape. Further elucidation of the aberrant molecular mechanisms that cause this acanthocytic state may possibly help to evaluate the pathological pathways leading to neurodegeneration.

  20. Graft-infiltrating host dendritic cells play a key role in organ transplant rejection

    Science.gov (United States)

    Zhuang, Quan; Liu, Quan; Divito, Sherrie J.; Zeng, Qiang; Yatim, Karim M.; Hughes, Andrew D.; Rojas-Canales, Darling M.; Nakao, A.; Shufesky, William J.; Williams, Amanda L.; Humar, Rishab; Hoffman, Rosemary A.; Shlomchik, Warren D.; Oberbarnscheidt, Martin H.; Lakkis, Fadi G.; Morelli, Adrian E.

    2016-01-01

    Successful engraftment of organ transplants has traditionally relied on preventing the activation of recipient (host) T cells. Once T-cell activation has occurred, however, stalling the rejection process becomes increasingly difficult, leading to graft failure. Here we demonstrate that graft-infiltrating, recipient (host) dendritic cells (DCs) play a key role in driving the rejection of transplanted organs by activated (effector) T cells. We show that donor DCs that accompany heart or kidney grafts are rapidly replaced by recipient DCs. The DCs originate from non-classical monocytes and form stable, cognate interactions with effector T cells in the graft. Eliminating recipient DCs reduces the proliferation and survival of graft-infiltrating T cells and abrogates ongoing rejection or rejection mediated by transferred effector T cells. Therefore, host DCs that infiltrate transplanted organs sustain the alloimmune response after T-cell activation has already occurred. Targeting these cells provides a means for preventing or treating rejection. PMID:27554168

  1. Graft-infiltrating host dendritic cells play a key role in organ transplant rejection.

    Science.gov (United States)

    Zhuang, Quan; Liu, Quan; Divito, Sherrie J; Zeng, Qiang; Yatim, Karim M; Hughes, Andrew D; Rojas-Canales, Darling M; Nakao, A; Shufesky, William J; Williams, Amanda L; Humar, Rishab; Hoffman, Rosemary A; Shlomchik, Warren D; Oberbarnscheidt, Martin H; Lakkis, Fadi G; Morelli, Adrian E

    2016-01-01

    Successful engraftment of organ transplants has traditionally relied on preventing the activation of recipient (host) T cells. Once T-cell activation has occurred, however, stalling the rejection process becomes increasingly difficult, leading to graft failure. Here we demonstrate that graft-infiltrating, recipient (host) dendritic cells (DCs) play a key role in driving the rejection of transplanted organs by activated (effector) T cells. We show that donor DCs that accompany heart or kidney grafts are rapidly replaced by recipient DCs. The DCs originate from non-classical monocytes and form stable, cognate interactions with effector T cells in the graft. Eliminating recipient DCs reduces the proliferation and survival of graft-infiltrating T cells and abrogates ongoing rejection or rejection mediated by transferred effector T cells. Therefore, host DCs that infiltrate transplanted organs sustain the alloimmune response after T-cell activation has already occurred. Targeting these cells provides a means for preventing or treating rejection.

  2. Graft-infiltrating host dendritic cells play a key role in organ transplant rejection.

    Science.gov (United States)

    Zhuang, Quan; Liu, Quan; Divito, Sherrie J; Zeng, Qiang; Yatim, Karim M; Hughes, Andrew D; Rojas-Canales, Darling M; Nakao, A; Shufesky, William J; Williams, Amanda L; Humar, Rishab; Hoffman, Rosemary A; Shlomchik, Warren D; Oberbarnscheidt, Martin H; Lakkis, Fadi G; Morelli, Adrian E

    2016-01-01

    Successful engraftment of organ transplants has traditionally relied on preventing the activation of recipient (host) T cells. Once T-cell activation has occurred, however, stalling the rejection process becomes increasingly difficult, leading to graft failure. Here we demonstrate that graft-infiltrating, recipient (host) dendritic cells (DCs) play a key role in driving the rejection of transplanted organs by activated (effector) T cells. We show that donor DCs that accompany heart or kidney grafts are rapidly replaced by recipient DCs. The DCs originate from non-classical monocytes and form stable, cognate interactions with effector T cells in the graft. Eliminating recipient DCs reduces the proliferation and survival of graft-infiltrating T cells and abrogates ongoing rejection or rejection mediated by transferred effector T cells. Therefore, host DCs that infiltrate transplanted organs sustain the alloimmune response after T-cell activation has already occurred. Targeting these cells provides a means for preventing or treating rejection. PMID:27554168

  3. Carcinoma cells misuse the host tissue damage response to invade the brain

    OpenAIRE

    Chuang, Han-Ning; van Rossum, Denise; Sieger, Dirk; Siam, Laila; Klemm, Florian; Bleckmann, Annalen; Bayerlová, Michaela; Farhat, Katja; Scheffel, Jörg; Schulz, Matthias; Dehghani, Faramarz; Stadelmann, Christine; Hanisch, Uwe-Karsten; Binder, Claudia; Pukrop, Tobias

    2013-01-01

    The metastatic colonization of the brain by carcinoma cells is still barely understood, in particular when considering interactions with the host tissue. The colonization comes with a substantial destruction of the surrounding host tissue. This leads to activation of damage responses by resident innate immune cells to protect, repair, and organize the wound healing, but may distract from tumoricidal actions. We recently demonstrated that microglia, innate immune cells of the CNS, assist carci...

  4. A Sequential Model of Host Cell Killing and Phagocytosis by Entamoeba histolytica

    OpenAIRE

    Adam Sateriale; Huston, Christopher D.

    2011-01-01

    The protozoan parasite Entamoeba histolytica is responsible for invasive intestinal and extraintestinal amebiasis. The virulence of Entamoeba histolytica is strongly correlated with the parasite's capacity to effectively kill and phagocytose host cells. The process by which host cells are killed and phagocytosed follows a sequential model of adherence, cell killing, initiation of phagocytosis, and engulfment. This paper presents recent advances in the cytolytic and phagocytic processes of Ent...

  5. Host parasite communications-Messages from helminths for the immune system: Parasite communication and cell-cell interactions.

    Science.gov (United States)

    Coakley, Gillian; Buck, Amy H; Maizels, Rick M

    2016-07-01

    Helminths are metazoan organisms many of which have evolved parasitic life styles dependent on sophisticated manipulation of the host environment. Most notably, they down-regulate host immune responses to ensure their own survival, by exporting a range of immuno-modulatory mediators that interact with host cells and tissues. While a number of secreted immunoregulatory parasite proteins have been defined, new work also points to the release of extracellular vesicles, or exosomes, that interact with and manipulate host gene expression. These recent results are discussed in the overall context of how helminths communicate effectively with the host organism. PMID:27297184

  6. Host parasite communications-Messages from helminths for the immune system: Parasite communication and cell-cell interactions.

    Science.gov (United States)

    Coakley, Gillian; Buck, Amy H; Maizels, Rick M

    2016-07-01

    Helminths are metazoan organisms many of which have evolved parasitic life styles dependent on sophisticated manipulation of the host environment. Most notably, they down-regulate host immune responses to ensure their own survival, by exporting a range of immuno-modulatory mediators that interact with host cells and tissues. While a number of secreted immunoregulatory parasite proteins have been defined, new work also points to the release of extracellular vesicles, or exosomes, that interact with and manipulate host gene expression. These recent results are discussed in the overall context of how helminths communicate effectively with the host organism.

  7. Failure to interact with Brd4 alters the ability of HPV16 E2 to regulate host genome expression and cellular movement.

    Science.gov (United States)

    Gauson, Elaine J; Wang, Xu; Dornan, Edward S; Herzyk, Pawel; Bristol, Molly; Morgan, Iain M

    2016-01-01

    The E2 protein of the carcinogen human papillomavirus 16 (HPV16) regulates replication and transcription of the viral genome in association with viral and cellular proteins. Our previous work demonstrated that E2 can regulate transcription from the host genome. E2 can activate transcription from adjacent promoters when located upstream using E2 DNA binding sequences and this activation is dependent upon the cellular protein Brd4; this report demonstrates that a Brd4 binding E2 mutant alters host genome expression differently from wild type E2. Of particular note is that highly down regulated genes are mostly not affected by failure to interact with Brd4 suggesting that the E2-Brd4 interaction is more responsible for the transcriptional activation of host genes rather than repression. Therefore failure to interact efficiently with Brd4, or altered levels of Brd4, would alter the ability of E2 to regulate the host genome and could contribute to determining the outcome of infection. PMID:26365679

  8. Alterations in integrin expression modulates invasion of pancreatic cancer cells.

    LENUS (Irish Health Repository)

    Walsh, Naomi

    2009-01-01

    BACKGROUND: Factors mediating the invasion of pancreatic cancer cells through the extracellular matrix (ECM) are not fully understood. METHODS: In this study, sub-populations of the human pancreatic cancer cell line, MiaPaCa-2 were established which displayed differences in invasion, adhesion, anoikis, anchorage-independent growth and integrin expression. RESULTS: Clone #3 displayed higher invasion with less adhesion, while Clone #8 was less invasive with increased adhesion to ECM proteins compared to MiaPaCa-2. Clone #8 was more sensitive to anoikis than Clone #3 and MiaPaCa-2, and displayed low colony-forming efficiency in an anchorage-independent growth assay. Integrins beta 1, alpha 5 and alpha 6 were over-expressed in Clone #8. Using small interfering RNA (siRNA), integrin beta1 knockdown in Clone #8 cells increased invasion through matrigel and fibronectin, increased motility, decreased adhesion and anoikis. Integrin alpha 5 and alpha 6 knockdown also resulted in increased motility, invasion through matrigel and decreased adhesion. CONCLUSION: Our results suggest that altered expression of integrins interacting with different extracellular matrixes may play a significant role in suppressing the aggressive invasive phenotype. Analysis of these clonal populations of MiaPaCa-2 provides a model for investigations into the invasive properties of pancreatic carcinoma.

  9. Withaferin a alters intermediate filament organization, cell shape and behavior.

    Directory of Open Access Journals (Sweden)

    Boris Grin

    Full Text Available Withaferin A (WFA is a steroidal lactone present in Withania somnifera which has been shown in vitro to bind to the intermediate filament protein, vimentin. Based upon its affinity for vimentin, it has been proposed that WFA can be used as an anti-tumor agent to target metastatic cells which up-regulate vimentin expression. We show that WFA treatment of human fibroblasts rapidly reorganizes vimentin intermediate filaments (VIF into a perinuclear aggregate. This reorganization is dose dependent and is accompanied by a change in cell shape, decreased motility and an increase in vimentin phosphorylation at serine-38. Furthermore, vimentin lacking cysteine-328, the proposed WFA binding site, remains sensitive to WFA demonstrating that this site is not required for its cellular effects. Using analytical ultracentrifugation, viscometry, electron microscopy and sedimentation assays we show that WFA has no effect on VIF assembly in vitro. Furthermore, WFA is not specific for vimentin as it disrupts the cellular organization and induces perinuclear aggregates of several other IF networks comprised of peripherin, neurofilament-triplet protein, and keratin. In cells co-expressing keratin IF and VIF, the former are significantly less sensitive to WFA with respect to inducing perinuclear aggregates. The organization of microtubules and actin/microfilaments is also affected by WFA. Microtubules become wavier and sparser and the number of stress fibers appears to increase. Following 24 hrs of exposure to doses of WFA that alter VIF organization and motility, cells undergo apoptosis. Lower doses of the drug do not kill cells but cause them to senesce. In light of our findings that WFA affects multiple IF systems, which are expressed in many tissues of the body, caution is warranted in its use as an anti-cancer agent, since it may have debilitating organism-wide effects.

  10. Galactose/N-acetylgalactosamine lectin: the coordinator of host cell killing

    Indian Academy of Sciences (India)

    Douglas R Boettner; Christopher Huston; William A Petri Jr

    2002-11-01

    Entamoeba histolytica is an enteric parasite that can kill host cells via a contact-dependent mechanism. This killing involves the amoebic surface protein referred to as the Gal/GalNAc lectin. The Gal/GalNAc lectin binds galactose and N-acetylgalactosamine allowing the adherence of amoebas to host cells. Involvement of the lectin in the pathogenesis of E. histolytica infection will be reviewed in this paper. The lectin has been shown to have very specific and substantial effects on adherence, cytotoxicity, and encystation. There is also possible involvement of the lectin in phagocytosis and caspase activation in host cells.

  11. Characterization of host lymphoid cells in antibody-facilitated bone marrow chimeras

    International Nuclear Information System (INIS)

    The authors have produced stable murine antibody-facilitated (AF) chimeras by the simultaneous injection of P1 bone marrow cells and anti-P2 monoclonal antibody into normal (unirradiated) adult (P1 X P2)F1 recipients. These AF chimeras are healthy, long-lived, and exhibit no overt signs of graft-versus-host disease. They are immunocompetent and tolerant of host, P2-encoded alloantigens. Donor cell engraftment and takeover, monitored by glucosephosphate isomerase isozyme patterns, is usually complete (greater than 95%) in the peripheral blood, bone marrow, and hemopoietic stem cell compartments of long-term (greater than 3 months posttransplantation) AF chimeras. The authors report here, however, that splenic, lymph node, and thymic leukocytes of AF chimeras represent donor/host chimeric populations. Spleen cell populations of AF chimeras exhibit substantial chimera-to-chimera variation in the preponderant residual host cell type(s) present. Interpretations of the implications of these findings are discussed

  12. Gut microbiome-derived metabolites modulate intestinal epithelial cell damage and mitigate graft-versus-host disease.

    Science.gov (United States)

    Mathewson, Nathan D; Jenq, Robert; Mathew, Anna V; Koenigsknecht, Mark; Hanash, Alan; Toubai, Tomomi; Oravecz-Wilson, Katherine; Wu, Shin-Rong; Sun, Yaping; Rossi, Corinne; Fujiwara, Hideaki; Byun, Jaeman; Shono, Yusuke; Lindemans, Caroline; Calafiore, Marco; Schmidt, Thomas C; Honda, Kenya; Young, Vincent B; Pennathur, Subramaniam; van den Brink, Marcel; Reddy, Pavan

    2016-05-01

    The effect of alterations in intestinal microbiota on microbial metabolites and on disease processes such as graft-versus-host disease (GVHD) is not known. Here we carried out an unbiased analysis to identify previously unidentified alterations in gastrointestinal microbiota-derived short-chain fatty acids (SCFAs) after allogeneic bone marrow transplant (allo-BMT). Alterations in the amount of only one SCFA, butyrate, were observed only in the intestinal tissue. The reduced butyrate in CD326(+) intestinal epithelial cells (IECs) after allo-BMT resulted in decreased histone acetylation, which was restored after local administration of exogenous butyrate. Butyrate restoration improved IEC junctional integrity, decreased apoptosis and mitigated GVHD. Furthermore, alteration of the indigenous microbiota with 17 rationally selected strains of high butyrate-producing Clostridia also decreased GVHD. These data demonstrate a heretofore unrecognized role of microbial metabolites and suggest that local and specific alteration of microbial metabolites has direct salutary effects on GVHD target tissues and can mitigate disease severity.

  13. Gut microbiome-derived metabolites modulate intestinal epithelial cell damage and mitigate graft-versus-host disease.

    Science.gov (United States)

    Mathewson, Nathan D; Jenq, Robert; Mathew, Anna V; Koenigsknecht, Mark; Hanash, Alan; Toubai, Tomomi; Oravecz-Wilson, Katherine; Wu, Shin-Rong; Sun, Yaping; Rossi, Corinne; Fujiwara, Hideaki; Byun, Jaeman; Shono, Yusuke; Lindemans, Caroline; Calafiore, Marco; Schmidt, Thomas C; Honda, Kenya; Young, Vincent B; Pennathur, Subramaniam; van den Brink, Marcel; Reddy, Pavan

    2016-05-01

    The effect of alterations in intestinal microbiota on microbial metabolites and on disease processes such as graft-versus-host disease (GVHD) is not known. Here we carried out an unbiased analysis to identify previously unidentified alterations in gastrointestinal microbiota-derived short-chain fatty acids (SCFAs) after allogeneic bone marrow transplant (allo-BMT). Alterations in the amount of only one SCFA, butyrate, were observed only in the intestinal tissue. The reduced butyrate in CD326(+) intestinal epithelial cells (IECs) after allo-BMT resulted in decreased histone acetylation, which was restored after local administration of exogenous butyrate. Butyrate restoration improved IEC junctional integrity, decreased apoptosis and mitigated GVHD. Furthermore, alteration of the indigenous microbiota with 17 rationally selected strains of high butyrate-producing Clostridia also decreased GVHD. These data demonstrate a heretofore unrecognized role of microbial metabolites and suggest that local and specific alteration of microbial metabolites has direct salutary effects on GVHD target tissues and can mitigate disease severity. PMID:26998764

  14. Host Range Expansion of Honey Bee Black Queen Cell Virus in the Bumble Bee, Bombus huntii

    Science.gov (United States)

    Honey bee viruses display a host range that is not restricted to their original host, European honey bees, Apis mellifera. Here we provide the first evidence that Black Queen Cell Virus (BQCV), one of the most prevalent honey bee viruses, can cause an infection in both laboratory-reared and field-co...

  15. Genome wide transcriptome analysis of dendritic cells identifies genes with altered expression in psoriasis.

    Directory of Open Access Journals (Sweden)

    Kata Filkor

    Full Text Available Activation of dendritic cells by different pathogens induces the secretion of proinflammatory mediators resulting in local inflammation. Importantly, innate immunity must be properly controlled, as its continuous activation leads to the development of chronic inflammatory diseases such as psoriasis. Lipopolysaccharide (LPS or peptidoglycan (PGN induced tolerance, a phenomenon of transient unresponsiveness of cells to repeated or prolonged stimulation, proved valuable model for the study of chronic inflammation. Thus, the aim of this study was the identification of the transcriptional diversity of primary human immature dendritic cells (iDCs upon PGN induced tolerance. Using SAGE-Seq approach, a tag-based transcriptome sequencing method, we investigated gene expression changes of primary human iDCs upon stimulation or restimulation with Staphylococcus aureus derived PGN, a widely used TLR2 ligand. Based on the expression pattern of the altered genes, we identified non-tolerizeable and tolerizeable genes. Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (Kegg analysis showed marked enrichment of immune-, cell cycle- and apoptosis related genes. In parallel to the marked induction of proinflammatory mediators, negative feedback regulators of innate immunity, such as TNFAIP3, TNFAIP8, Tyro3 and Mer are markedly downregulated in tolerant cells. We also demonstrate, that the expression pattern of TNFAIP3 and TNFAIP8 is altered in both lesional, and non-lesional skin of psoriatic patients. Finally, we show that pretreatment of immature dendritic cells with anti-TNF-α inhibits the expression of IL-6 and CCL1 in tolerant iDCs and partially releases the suppression of TNFAIP8. Our findings suggest that after PGN stimulation/restimulation the host cell utilizes different mechanisms in order to maintain critical balance between inflammation and tolerance. Importantly, the transcriptome sequencing of stimulated/restimulated iDCs identified

  16. Antibiotic-induced dysbiosis alters host-bacterial interactions and leads to colonic sensory and motor changes in mice.

    Science.gov (United States)

    Aguilera, M; Cerdà-Cuéllar, M; Martínez, V

    2015-01-01

    Alterations in the composition of the commensal microbiota (dysbiosis) seem to be a pathogenic component of functional gastrointestinal disorders, mainly irritable bowel syndrome (IBS), and might participate in the secretomotor and sensory alterations observed in these patients.We determined if a state antibiotics-induced intestinal dysbiosis is able to modify colonic pain-related and motor responses and characterized the neuro-immune mechanisms implicated in mice. A 2-week antibiotics treatment induced a colonic dysbiosis (increments in Bacteroides spp, Clostridium coccoides and Lactobacillus spp and reduction in Bifidobacterium spp). Bacterial adherence was not affected. Dysbiosis was associated with increased levels of secretory-IgA, up-regulation of the antimicrobial lectin RegIIIγ, and toll-like receptors (TLR) 4 and 7 and down-regulation of the antimicrobial-peptide Resistin-Like Molecule-β and TLR5. Dysbiotic mice showed less goblet cells, without changes in the thickness of the mucus layer. Neither macroscopical nor microscopical signs of inflammation were observed. In dysbiotic mice, expression of the cannabinoid receptor 2 was up-regulated, while the cannabinoid 1 and the mu-opioid receptors were down-regulated. In antibiotic-treated mice, visceral pain-related responses elicited by intraperitoneal acetic acid or intracolonic capsaicin were significantly attenuated. Colonic contractility was enhanced during dysbiosis. Intestinal dysbiosis induce changes in the innate intestinal immune system and modulate the expression of pain-related sensory systems, an effect associated with a reduction in visceral pain-related responses. Commensal microbiota modulates gut neuro-immune sensory systems, leading to functional changes, at least as it relates to viscerosensitivity. Similar mechanisms might explain the beneficial effects of antibiotics or certain probiotics in the treatment of IBS. PMID:25531553

  17. DMPD: The role of type I interferon production by dendritic cells in host defense. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17544561 The role of type I interferon production by dendritic cells in host defens...tml) (.csml) Show The role of type I interferon production by dendritic cells in host defense. PubmedID 1754...4561 Title The role of type I interferon production by dendritic cells in host de

  18. Bacterial cell-cell communication in the host via RRNPP peptide-binding regulators

    Directory of Open Access Journals (Sweden)

    David ePerez-Pascual

    2016-05-01

    Full Text Available Human microbiomes are composed of complex and dense bacterial consortia. In these environments, bacteria are able to react quickly to change by coordinating their gene expression at the population level via small signaling molecules. In Gram-positive bacteria, cell-cell communication is mostly mediated by peptides that are released into the extracellular environment. Cell-cell communication based on these peptides is especially widespread in the group Firmicutes, in which they regulate a wide array of biological processes, including functions related to host-microbe interactions. Among the different agents of communication, the RRNPP family of cytoplasmic transcriptional regulators, together with their cognate re-internalized signaling peptides, represents a group of emerging importance. RRNPP members that have been studied so far are found mainly in species of bacilli, streptococci, and enterococci. These bacteria are characterized as both human commensal and pathogenic, and share different niches in the human body with other microorganisms. The goal of this mini-review is to present the current state of research on the biological relevance of RRNPP mechanisms in the context of the host, highlighting their specific roles in commensalism or virulence.

  19. Staphylococcus aureus produces membrane-derived vesicles that induce host cell death.

    Directory of Open Access Journals (Sweden)

    Mamata Gurung

    Full Text Available Gram-negative bacteria produce outer membrane vesicles that play a role in the delivery of virulence factors to host cells. However, little is known about the membrane-derived vesicles (MVs produced by gram-positive bacteria. The present study examined the production of MVs from Staphylococcus aureus and investigated the delivery of MVs to host cells and subsequent cytotoxicity. Four S. aureus strains tested, two type strains and two clinical isolates, produced spherical nanovesicles during in vitro culture. MVs were also produced during in vivo infection of a clinical S. aureus isolate in a mouse pneumonia model. Proteomic analysis showed that 143 different proteins were identified in the S. aureus-derived MVs. S. aureus MVs were interacted with the plasma membrane of host cells via a cholesterol-rich membrane microdomain and then delivered their component protein A to host cells within 30 min. Intact S. aureus MVs induced apoptosis of HEp-2 cells in a dose-dependent manner, whereas lysed MVs neither delivered their component into the cytosol of host cells nor induced cytotoxicity. In conclusion, this study is the first report that S. aureus MVs are an important vehicle for delivery of bacterial effector molecules to host cells.

  20. HIV-host interactome revealed directly from infected cells.

    Science.gov (United States)

    Luo, Yang; Jacobs, Erica Y; Greco, Todd M; Mohammed, Kevin D; Tong, Tommy; Keegan, Sarah; Binley, James M; Cristea, Ileana M; Fenyö, David; Rout, Michael P; Chait, Brian T; Muesing, Mark A

    2016-01-01

    Although genetically compact, HIV-1 commandeers vast arrays of cellular machinery to sustain and protect it during cycles of viral outgrowth. Transposon-mediated saturation linker scanning mutagenesis was used to isolate fully replication-competent viruses harbouring a potent foreign epitope tag. Using these viral isolates, we performed differential isotopic labelling and affinity-capture mass spectrometric analyses on samples obtained from cultures of human lymphocytes to classify the vicinal interactomes of the viral Env and Vif proteins as they occur during natural infection. Importantly, interacting proteins were recovered without bias, regardless of their potential for positive, negative or neutral impact on viral replication. We identified specific host associations made with trimerized Env during its biosynthesis, at virological synapses, with innate immune effectors (such as HLA-E) and with certain cellular signalling pathways (for example, Notch1). We also defined Vif associations with host proteins involved in the control of nuclear transcription and nucleoside biosynthesis as well as those interacting stably or transiently with the cytoplasmic protein degradation apparatus. Our approach is broadly applicable to elucidating pathogen-host interactomes, providing high-certainty identification of interactors by their direct access during cycling infection. Understanding the pathophysiological consequences of these associations is likely to provide strategic targets for antiviral intervention. PMID:27572969

  1. Interleukin-7 Modulates Anti-Tumor CD8+ T Cell Responses via Its Action on Host Cells

    Science.gov (United States)

    Deiser, Katrin; Stoycheva, Diana; Bank, Ute; Blankenstein, Thomas; Schüler, Thomas

    2016-01-01

    The adoptive transfer of antigen-specific CD8+ T cells is a promising approach for the treatment of chronic viral and malignant diseases. In order to improve adoptive T cell therapy (ATT) of cancer, recent strategies aim at the antibody-based blockade of immunosuppressive signaling pathways in CD8+ T cells. Alternatively, adjuvant effects of immunostimulatory cytokines might be exploited to improve therapeutic CD8+ T cell responses. For example, Interleukin-7 (IL-7) is a potent growth, activation and survival factor for CD8+ T cells that can be used to improve virus- and tumor-specific CD8+ T cell responses. Although direct IL-7 effects on CD8+ T cells were studied extensively in numerous models, the contribution of IL-7 receptor-competent (IL-7R+) host cells remained unclear. In the current study we provide evidence that CD8+ T cell-mediated tumor rejection in response to recombinant IL-7 (rIL-7) therapy is strictly dependent on IL-7R+ host cells. On the contrary, CD8+ T cell expansion is independent of host IL-7R expression. If, however, rIL-7 therapy and peptide vaccination are combined, host IL-7R signaling is crucial for CD8+ T cell expansion. Unexpectedly, maximum CD8+ T cell expansion relies mainly on IL-7R signaling in non-hematopoietic host cells, similar to the massive accumulation of dendritic cells and granulocytes. In summary, we provide evidence that IL-7R+ host cells are major targets of rIL-7 that modulate therapeutic CD8+ T cell responses and the outcome of rIL-7-assisted ATT. This knowledge may have important implications for the design and optimization of clinical ATT protocols. PMID:27447484

  2. Interleukin-7 Modulates Anti-Tumor CD8+ T Cell Responses via Its Action on Host Cells.

    Directory of Open Access Journals (Sweden)

    Katrin Deiser

    Full Text Available The adoptive transfer of antigen-specific CD8+ T cells is a promising approach for the treatment of chronic viral and malignant diseases. In order to improve adoptive T cell therapy (ATT of cancer, recent strategies aim at the antibody-based blockade of immunosuppressive signaling pathways in CD8+ T cells. Alternatively, adjuvant effects of immunostimulatory cytokines might be exploited to improve therapeutic CD8+ T cell responses. For example, Interleukin-7 (IL-7 is a potent growth, activation and survival factor for CD8+ T cells that can be used to improve virus- and tumor-specific CD8+ T cell responses. Although direct IL-7 effects on CD8+ T cells were studied extensively in numerous models, the contribution of IL-7 receptor-competent (IL-7R+ host cells remained unclear. In the current study we provide evidence that CD8+ T cell-mediated tumor rejection in response to recombinant IL-7 (rIL-7 therapy is strictly dependent on IL-7R+ host cells. On the contrary, CD8+ T cell expansion is independent of host IL-7R expression. If, however, rIL-7 therapy and peptide vaccination are combined, host IL-7R signaling is crucial for CD8+ T cell expansion. Unexpectedly, maximum CD8+ T cell expansion relies mainly on IL-7R signaling in non-hematopoietic host cells, similar to the massive accumulation of dendritic cells and granulocytes. In summary, we provide evidence that IL-7R+ host cells are major targets of rIL-7 that modulate therapeutic CD8+ T cell responses and the outcome of rIL-7-assisted ATT. This knowledge may have important implications for the design and optimization of clinical ATT protocols.

  3. Interleukin-7 Modulates Anti-Tumor CD8+ T Cell Responses via Its Action on Host Cells.

    Science.gov (United States)

    Deiser, Katrin; Stoycheva, Diana; Bank, Ute; Blankenstein, Thomas; Schüler, Thomas

    2016-01-01

    The adoptive transfer of antigen-specific CD8+ T cells is a promising approach for the treatment of chronic viral and malignant diseases. In order to improve adoptive T cell therapy (ATT) of cancer, recent strategies aim at the antibody-based blockade of immunosuppressive signaling pathways in CD8+ T cells. Alternatively, adjuvant effects of immunostimulatory cytokines might be exploited to improve therapeutic CD8+ T cell responses. For example, Interleukin-7 (IL-7) is a potent growth, activation and survival factor for CD8+ T cells that can be used to improve virus- and tumor-specific CD8+ T cell responses. Although direct IL-7 effects on CD8+ T cells were studied extensively in numerous models, the contribution of IL-7 receptor-competent (IL-7R+) host cells remained unclear. In the current study we provide evidence that CD8+ T cell-mediated tumor rejection in response to recombinant IL-7 (rIL-7) therapy is strictly dependent on IL-7R+ host cells. On the contrary, CD8+ T cell expansion is independent of host IL-7R expression. If, however, rIL-7 therapy and peptide vaccination are combined, host IL-7R signaling is crucial for CD8+ T cell expansion. Unexpectedly, maximum CD8+ T cell expansion relies mainly on IL-7R signaling in non-hematopoietic host cells, similar to the massive accumulation of dendritic cells and granulocytes. In summary, we provide evidence that IL-7R+ host cells are major targets of rIL-7 that modulate therapeutic CD8+ T cell responses and the outcome of rIL-7-assisted ATT. This knowledge may have important implications for the design and optimization of clinical ATT protocols.

  4. Heterologously expressed Staphylococcus aureus fibronectin-binding proteins are sufficient for invasion of host cells

    NARCIS (Netherlands)

    Sinha, B; Francois, P; Que, Y A; Hussain, M; Heilmann, C; Moreillon, P; Lew, D; Krause, K H; Peters, G; Herrmann, M

    2000-01-01

    Staphylococcus aureus invasion of mammalian cells, including epithelial, endothelial, and fibroblastic cells, critically depends on fibronectin bridging between S. aureus fibronectin-binding proteins (FnBPs) and the host fibronectin receptor integrin alpha(5)beta(1) (B. Sinha et al., Cell. Microbiol

  5. Adhesion defective BHK cell mutant has cell surface heparan sulfate proteoglycan of altered properties

    DEFF Research Database (Denmark)

    Couchman, J R; Austria, R; Woods, A;

    1988-01-01

    sulfation, reduced affinity for fibronectin and decreased half-life on the cell surface when compared to the normal counterpart. Our conclusions based on this data are that these altered properties may, in part, account for the adhesion defect in the ricin-resistant mutant. Whether this results from......In the light of accumulating data that implicate cell surface heparan sulfate proteoglycans (HSPGs) with a role in cell interactions with extracellular matrix molecules such as fibronectin, we have compared the properties of these molecules in wild-type BHK cells and an adhesion-defective ricin......-resistant mutant (RicR14). Our results showed that the mutant, unlike BHK cells, cannot form focal adhesions when adherent to planar substrates in the presence of serum. Furthermore, while both cell lines possess similar amounts of cell surface HSPG with hydrophobic properties, that of RicR14 cells had decreased...

  6. Cell Transformation and Proteome Alteration in QSG7701 Cells Transfected with Hepatitis C Virus Non-structural Protein 3

    Institute of Scientific and Technical Information of China (English)

    Qiongqiong HE; Deyun FENG; Ruixue CHENG; Zhuchu CHEN; Xuxian XIAO; Zhiqiang XIAO; Cui LI; Bo LI; Pengfei ZHANG; Hui ZHENG

    2007-01-01

    Persistent hepatitis C virus (HCV) infection can cause liver cirrhosis and hepatocellular carcinoma. Non-structural protein 3 (NS3), an important part of HCV, has been implicated in the life cycle of the virus and interacts with host cellular proteins. In this study, we investigated the effect of NS3 protein on cell tranformation and related protein alteration in human hepatocyte QSG7701 cells. The results indicated that stable expression of the NS3 protein in QSG7701 cells induced transformed characters with reduced population doubling time, anchorage-independent growth and tumor development. Fifteen differentially-expressed proteins were separated and identified using 2-D electrophoresis and matrix-assisted laser desorption ionization-time of flight mass spectrometry. Western blot analysis confirmed that the increase of phospho-p44/42 and phospho-p38 proteins was associated with transformed cells. These results supported the view that HCV NS3 protein plays a transforming role and provided some clues to elucidate the carcinogenesis mechanism of HCV-related hepatocellular carcinoma.

  7. Noncoding RNA small nucleolar RNA host gene 1 promote cell proliferation in nonsmall cell lung cancer

    Directory of Open Access Journals (Sweden)

    J You

    2014-01-01

    Full Text Available Background: Nonsmall cell lung cancer (NSCLC is the major cause of cancer death worldwide. Increasing evidence shows that noncoding RNAs (ncRNAs are widely involved in the development and progression of NSCLC. ncRNA small nucleolar RNA host gene 1 (SNHG1 has not been studied in cancer, especially its role in lung cancer remains unknown. Our studies were designed to investigate the expression and biological significance of SNHG1 in lung cancer. SNHG1 may be a novel ncRNA in early diagnosis in lung cancer. Methods: Noncoding RNA SNHG1 expression in 7 lung cancer cell lines was measured by quantitative real-time polymerase chain reaction. RNA interference approaches were used to find the biological functions of SNHG1. The effect of SNHG1 on proliferation was evaluated by cell count and crystal violet stains. Results: Noncoding RNA SNHG1 expression was significantly upregulated in lung cancer cells when compared with normal bronchial epithelial cells. In addition, in vitro assays our results indicated that knockdown of SNHG1 inhibited cell proliferation. Conclusions: Our data indicated that ncRNA SNHG1 is significantly upregulated in NSCLC cell lines and may represent a new biomarker and a potential therapeutic target for NSCLC intervention.

  8. Altered cell cycle regulation helps stem-like carcinoma cells resist apoptosis

    OpenAIRE

    Dalton Stephen; Chappell James

    2010-01-01

    Abstract Reemergence of carcinomas following chemotherapy and/or radiotherapy is not well understood, but a recent study in BMC Cancer suggests that resistance to apoptosis resulting from altered cell cycle regulation is crucial. See research article: http://biomedcentral.com/1471-2407/10/166

  9. Altered expression of epithelial cell surface glycoconjugates and intermediate filaments at the margins of mucosal wounds

    DEFF Research Database (Denmark)

    Dabelsteen, Erik; Grøn, B; Mandel, U;

    1998-01-01

    Alterations in cell to cell adhesion are necessary to enable the type of cell movements that are associated with epithelial wound healing and malignant invasion. Several studies of transformed cells have related epithelial cell movement to changes in the cell surface expression of the carbohydrat...

  10. Trypanosoma cruzi extracellular amastigotes and host cell signaling: more pieces to the puzzle

    Directory of Open Access Journals (Sweden)

    Éden Ramalho Ferreira

    2012-11-01

    Full Text Available Among the different infective stages that Trypanosoma cruzi employs to invade cells, extracellular amastigotes have recently gained attention by our group. This is true primarily because these amastigotes are able to infect cultured cells and animals, establishing a sustainable infective cycle. Extracellular amastigotes are thus an excellent means of adaptation and survival for T. cruzi, whose different infective stages each utilize unique mechanisms for attachment and penetration. Here we discuss some features of host cell invasion by extracellular amastigotes and the associated host cell signaling events that occur as part of the process.

  11. Bordetella pertussis naturally occurring isolates with altered lipooligosaccharide structure fail to fully mature human dendritic cells.

    Science.gov (United States)

    Brummelman, Jolanda; Veerman, Rosanne E; Hamstra, Hendrik Jan; Deuss, Anna J M; Schuijt, Tim J; Sloots, Arjen; Kuipers, Betsy; van Els, Cécile A C M; van der Ley, Peter; Mooi, Frits R; Han, Wanda G H; Pinelli, Elena

    2015-01-01

    Bordetella pertussis is a Gram-negative bacterium and the causative agent of whooping cough. Despite high vaccination coverage, outbreaks are being increasingly reported worldwide. Possible explanations include adaptation of this pathogen, which may interfere with recognition by the innate immune system. Here, we describe innate immune recognition and responses to different B. pertussis clinical isolates. By using HEK-Blue cells transfected with different pattern recognition receptors, we found that 3 out of 19 clinical isolates failed to activate Toll-like receptor 4 (TLR4). These findings were confirmed by using the monocytic MM6 cell line. Although incubation with high concentrations of these 3 strains resulted in significant activation of the MM6 cells, it was found to occur mainly through interaction with TLR2 and not through TLR4. When using live bacteria, these 3 strains also failed to activate TLR4 on HEK-Blue cells, and activation of MM6 cells or human monocyte-derived dendritic cells was significantly lower than activation induced by the other 16 strains. Mass spectrum analysis of the lipid A moieties from these 3 strains indicated an altered structure of this molecule. Gene sequence analysis revealed mutations in genes involved in lipid A synthesis. Findings from this study indicate that B. pertussis isolates that do not activate TLR4 occur naturally and that this phenotype may give this bacterium an advantage in tempering the innate immune response and establishing infection. Knowledge on the strategies used by this pathogen in evading the host immune response is essential for the improvement of current vaccines or for the development of new ones.

  12. Trichomonas vaginalis exosomes deliver cargo to host cells and mediate host∶parasite interactions.

    Directory of Open Access Journals (Sweden)

    Olivia Twu

    Full Text Available Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogential tract where it remains extracellular and adheres to epithelial cells. Infections range from asymptomatic to highly inflammatory, depending on the host and the parasite strain. Here, we use a combination of methodologies including cell fractionation, immunofluorescence and electron microscopy, RNA, proteomic and cytokine analyses and cell adherence assays to examine pathogenic properties of T. vaginalis. We have found that T.vaginalis produces and secretes microvesicles with physical and biochemical properties similar to mammalian exosomes. The parasite-derived exosomes are characterized by the presence of RNA and core, conserved exosomal proteins as well as parasite-specific proteins. We demonstrate that T. vaginalis exosomes fuse with and deliver their contents to host cells and modulate host cell immune responses. Moreover, exosomes from highly adherent parasite strains increase the adherence of poorly adherent parasites to vaginal and prostate epithelial cells. In contrast, exosomes from poorly adherent strains had no measurable effect on parasite adherence. Exosomes from parasite strains that preferentially bind prostate cells increased binding of parasites to these cells relative to vaginal cells. In addition to establishing that parasite exosomes act to modulate host∶parasite interactions, these studies are the first to reveal a potential role for exosomes in promoting parasite∶parasite communication and host cell colonization.

  13. Host Cell Factors in Filovirus Entry: Novel Players, New Insights

    Directory of Open Access Journals (Sweden)

    Stefan Pöhlmann

    2012-11-01

    Full Text Available Filoviruses cause severe hemorrhagic fever in humans with high case-fatality rates. The cellular factors exploited by filoviruses for their spread constitute potential targets for intervention, but are incompletely defined. The viral glycoprotein (GP mediates filovirus entry into host cells. Recent studies revealed important insights into the host cell molecules engaged by GP for cellular entry. The binding of GP to cellular lectins was found to concentrate virions onto susceptible cells and might contribute to the early and sustained infection of macrophages and dendritic cells, important viral targets. Tyrosine kinase receptors were shown to promote macropinocytic uptake of filoviruses into a subset of susceptible cells without binding to GP, while interactions between GP and human T cell Ig mucin 1 (TIM-1 might contribute to filovirus infection of mucosal epithelial cells. Moreover, GP engagement of the cholesterol transporter Niemann-Pick C1 was demonstrated to be essential for GP-mediated fusion of the viral envelope with a host cell membrane. Finally, mutagenic and structural analyses defined GP domains which interact with these host cell factors. Here, we will review the recent progress in elucidating the molecular interactions underlying filovirus entry and discuss their implications for our understanding of the viral cell tropism.

  14. Analysis of host- and strain-dependent cell death responses during infectious salmon anemia virus infection in vitro

    Directory of Open Access Journals (Sweden)

    Mjaaland Siri

    2009-07-01

    Full Text Available Abstract Background Infectious salmon anemia virus (ISAV is an aquatic orthomyxovirus and the causative agent of infectious salmon anemia (ISA, a disease of great importance in the Atlantic salmon farming industry. In vitro, ISAV infection causes cytophatic effect (CPE in cell lines from Atlantic salmon, leading to rounding and finally detachment of the cells from the substratum. In this study, we investigated the mode of cell death during in vitro ISAV infection in different Atlantic salmon cell lines, using four ISAV strains causing different mortality in vivo. Results The results show that caspase 3/7 activity increased during the course of infection in ASK and SHK-1 cells, infected cells showed increased surface expression of phosphatidylserine and increased PI uptake, compared to mock infected cells; and morphological alterations of the mitochondria were observed. Expression analysis of immune relevant genes revealed no correlation between in vivo mortality and expression, but good correlation in expression of interferon genes. Conclusion Results from this study indicate that there is both strain and cell type dependent differences in the virus-host interaction during ISAV infection. This is important to bear in mind when extrapolating in vitro findings to the in vivo situation.

  15. Molecular and Cellular Mechanisms Involved in the Trypanosoma cruzi/Host Cell Interplay

    Science.gov (United States)

    Romano, Patricia Silvia; Cueto, Juan Agustín; Casassa, Ana Florencia; Vanrell, María Cristina; Gottlieb, Roberta A.; Colombo, María Isabel

    2013-01-01

    Summary The protozoan parasite Trypanosoma cruzi has a complex bi-ological cycle that involves vertebrate and invertebrate hosts. In mammals, the infective trypomastigote form of this parasite can invade several cell types by exploiting phagocytic-like or non-phagocytic mechanisms depending on the class of cell involved. Morphological studies showed that when trypomastigotes contact macrophages, they induce the formation of plasma membrane protrusions that differ from the canonical phagocytosis that occurs in the case of noninfective epimastigotes. In contrast, when trypomastigotes infect epithelial or muscle cells, the cell surface is minimally modified, suggesting the induction of a different class of process. Lysosomal-dependent or -independent T. cruzi invasion of host cells are two different models that describe the molecular and cellular events activated during parasite entry into nonphagocytic cells. In this context, we have previously shown that induction of autophagy in host cells before infection favors T. cruzi invasion. Furthermore, we demonstrate that autophagosomes and the autophagosomal protein LC3 are recruited to the T. cruzi entry sites and that the newly formed T. cruzi parasitophorous vacuole has characteristics of an autophagolysosome. This review summarizes the current knowledge of the molecular and cellular mechanisms of T. cruzi invasion in nonphagocytic cells. Based on our findings, we propose a new model in which T. cruzi takes advantage of the up-regulation of autophagy during starvation to increase its successful colonization of host cells. PMID:22454195

  16. Host cell proteins in biotechnology-derived products: A risk assessment framework.

    Science.gov (United States)

    de Zafra, Christina L Zuch; Quarmby, Valerie; Francissen, Kathleen; Vanderlaan, Martin; Zhu-Shimoni, Judith

    2015-11-01

    To manufacture biotechnology products, mammalian or bacterial cells are engineered for the production of recombinant therapeutic human proteins including monoclonal antibodies. Host cells synthesize an entire repertoire of proteins which are essential for their own function and survival. Biotechnology manufacturing processes are designed to produce recombinant therapeutics with a very high degree of purity. While there is typically a low residual level of host cell protein in the final drug product, under some circumstances a host cell protein(s) may copurify with the therapeutic protein and, if it is not detected and removed, it may become an unintended component of the final product. The purpose of this article is to enumerate and discuss factors to be considered in an assessment of risk of residual host cell protein(s) detected and identified in the drug product. The consideration of these factors and their relative ranking will lead to an overall risk assessment that informs decision-making around how to control the levels of host cell proteins.

  17. A gene-alteration profile of human lung cancer cell lines

    OpenAIRE

    R. Blanco; Iwakawa, R.; Tang, M; Kohno, T.; Angulo, B; Pio, R. (Rubén); Montuenga, L M; Minna, J D; Yokota, J; Sanchez-Cespedes, M.

    2009-01-01

    ABSTRACT: Aberrant proteins encoded from genes altered in tumors drive cancer development and may also be therapeutic targets. Here we derived a comprehensive gene-alteration profile of lung cancer cell lines. We tested 17 genes in a panel of 88 lung cancer cell lines and found the rates of alteration to be higher than previously thought. Nearly all cells feature inactivation at TP53 and CDKN2A or RB1, whereas BRAF, MET, ERBB2, and NRAS alterations were infrequent. A p...

  18. Transcriptome Analysis of HepG2 Cells Expressing ORF3 from Swine Hepatitis E Virus to Determine the Effects of ORF3 on Host Cells

    Directory of Open Access Journals (Sweden)

    Kailian Xu

    2016-01-01

    Full Text Available Hepatitis E virus- (HEV- mediated hepatitis has become a global public health problem. An important regulatory protein of HEV, ORF3, influences multiple signal pathways in host cells. In this study, to investigate the function of ORF3 from the swine form of HEV (SHEV, high-throughput RNA-Seq-based screening was performed to identify the differentially expressed genes in ORF3-expressing HepG2 cells. The results were validated with quantitative real-time PCR and gene ontology was employed to assign differentially expressed genes to functional categories. The results indicated that, in the established ORF3-expressing HepG2 cells, the mRNA levels of CLDN6, YLPM1, APOC3, NLRP1, SCARA3, FGA, FGG, FGB, and FREM1 were upregulated, whereas the mRNA levels of SLC2A3, DKK1, BPIFB2, and PTGR1 were downregulated. The deregulated expression of CLDN6 and FREM1 might contribute to changes in integral membrane protein and basement membrane protein expression, expression changes for NLRP1 might affect the apoptosis of HepG2 cells, and the altered expression of APOC3, SCARA3, and DKK1 may affect lipid metabolism in HepG2 cells. In conclusion, ORF3 plays a functional role in virus-cell interactions by affecting the expression of integral membrane protein and basement membrane proteins and by altering the process of apoptosis and lipid metabolism in host cells. These findings provide important insight into the pathogenic mechanism of HEV.

  19. Transcriptome Analysis of HepG2 Cells Expressing ORF3 from Swine Hepatitis E Virus to Determine the Effects of ORF3 on Host Cells

    Science.gov (United States)

    Guo, Shiyu; Zhao, Tianjing; Zhu, Huapei; Jiao, Hanwei; Shi, Qiaoyun; Pang, Feng; Li, Yaying; Li, Guohua; Peng, Dongmei; Nie, Xin; Wu, Kebang; Du, Li; Cui, Ke

    2016-01-01

    Hepatitis E virus- (HEV-) mediated hepatitis has become a global public health problem. An important regulatory protein of HEV, ORF3, influences multiple signal pathways in host cells. In this study, to investigate the function of ORF3 from the swine form of HEV (SHEV), high-throughput RNA-Seq-based screening was performed to identify the differentially expressed genes in ORF3-expressing HepG2 cells. The results were validated with quantitative real-time PCR and gene ontology was employed to assign differentially expressed genes to functional categories. The results indicated that, in the established ORF3-expressing HepG2 cells, the mRNA levels of CLDN6, YLPM1, APOC3, NLRP1, SCARA3, FGA, FGG, FGB, and FREM1 were upregulated, whereas the mRNA levels of SLC2A3, DKK1, BPIFB2, and PTGR1 were downregulated. The deregulated expression of CLDN6 and FREM1 might contribute to changes in integral membrane protein and basement membrane protein expression, expression changes for NLRP1 might affect the apoptosis of HepG2 cells, and the altered expression of APOC3, SCARA3, and DKK1 may affect lipid metabolism in HepG2 cells. In conclusion, ORF3 plays a functional role in virus-cell interactions by affecting the expression of integral membrane protein and basement membrane proteins and by altering the process of apoptosis and lipid metabolism in host cells. These findings provide important insight into the pathogenic mechanism of HEV. PMID:27648443

  20. A host-parasite model for a two-type cell population

    CERN Document Server

    Alsmeyer, Gerold

    2012-01-01

    A host-parasite model is considered for a population of cells that can be of two types, A or B, and exhibits unilateral reproduction: while a B-cell always splits into two cells of the same type, the two daughter cells of an A-cell can be of any type. The random mechanism that describes how parasites within a cell multiply and are then shared into the daughter cells is allowed to depend on the hosting mother cell as well as its daughter cells. Focusing on the subpopulation of A-cells and its parasites, the model differs from the single-type model recently studied by Bansaye (2008) in that the sharing mechanism may be biased towards one of the two types. Main results are concerned with the nonextinctive case and provide information on the behavior, as $n\\to\\infty$, of the number A-parasites in generation n and the relative proportion of A- and B-cells in this generation which host a given number of parasites. As in (Bansaye,2008), proofs will make use of a so-called random cell line which, when conditioned to ...

  1. The Vibrio parahaemolyticus Type III Secretion Systems manipulate host cell MAPK for critical steps in pathogenesis.

    LENUS (Irish Health Repository)

    Matlawska-Wasowska, Ksenia

    2010-12-01

    Vibrio parahaemolyticus is a food-borne pathogen causing inflammation of the gastrointestinal epithelium. Pathogenic strains of this bacterium possess two Type III Secretion Systems (TTSS) that deliver effector proteins into host cells. In order to better understand human host cell responses to V. parahaemolyticus, the modulation of Mitogen Activated Protein Kinase (MAPK) activation in epithelial cells by an O3:K6 clinical isolate, RIMD2210633, was investigated. The importance of MAPK activation for the ability of the bacterium to be cytotoxic and to induce secretion of Interleukin-8 (IL-8) was determined.

  2. In Vitro Exposure of Harbor Seal Immune Cells to Aroclor 1260 Alters Phocine Distemper Virus Replication.

    Science.gov (United States)

    Bogomolni, Andrea; Frasca, Salvatore; Levin, Milton; Matassa, Keith; Nielsen, Ole; Waring, Gordon; De Guise, Sylvain

    2016-01-01

    In the last 30 years, several large-scale marine mammal mortality events have occurred, often in close association with highly polluted regions, leading to suspicions that contaminant-induced immunosuppression contributed to these epizootics. Some of these recent events also identified morbillivirus as a cause of or contributor to death. The role of contaminant exposures regarding morbillivirus mortality is still unclear. The results of this study aimed to address the potential for a mixture of polychlorinated biphenyls (PCBs), specifically Aroclor 1260, to alter harbor seal T-lymphocyte proliferation and to assess if exposure resulted in increased likelihood of phocine distemper virus (PDV USA 2006) to infect susceptible seals in an in vitro system. Exposure of peripheral blood mononuclear cells to Aroclor 1260 did not significantly alter lymphocyte proliferation (1, 5, 10, and 20 ppm). However, using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), lymphocytes exposed to 20 ppm Aroclor 1260 exhibited a significant decrease in PDV replication at day 7 and a significant increase at day 11 compared with unexposed control cells. Similar and significant differences were apparent on exposure to Aroclor 1260 in monocytes and supernatant. The results here indicate that in harbor seals, Aroclor 1260 exposure results in a decrease in virus early during infection and an increase during late infection. The consequences of this contaminant-induced infection pattern in a highly susceptible host could result in a greater potential for systemic infection with greater viral load, which could explain the correlative findings seen in wild populations exposed to a range of persistent contaminants that suffer from morbillivirus epizootics. PMID:26142119

  3. Proteomic analysis of the action of the Mycobacterium ulcerans toxin mycolactone: targeting host cells cytoskeleton and collagen.

    Directory of Open Access Journals (Sweden)

    José B Gama

    2014-08-01

    Full Text Available Buruli ulcer (BU is a neglected tropical disease caused by Mycobacterium ulcerans. The tissue damage characteristic of BU lesions is known to be driven by the secretion of the potent lipidic exotoxin mycolactone. However, the molecular action of mycolactone on host cell biology mediating cytopathogenesis is not fully understood. Here we applied two-dimensional electrophoresis (2-DE to identify the mechanisms of mycolactone's cellular action in the L929 mouse fibroblast proteome. This revealed 20 changed spots corresponding to 18 proteins which were clustered mainly into cytoskeleton-related proteins (Dync1i2, Cfl1, Crmp2, Actg1, Stmn1 and collagen biosynthesis enzymes (Plod1, Plod3, P4ha1. In line with cytoskeleton conformational disarrangements that are observed by immunofluorescence, we found several regulators and constituents of both actin- and tubulin-cytoskeleton affected upon exposure to the toxin, providing a novel molecular basis for the effect of mycolactone. Consistent with these cytoskeleton-related alterations, accumulation of autophagosomes as well as an increased protein ubiquitination were observed in mycolactone-treated cells. In vivo analyses in a BU mouse model revealed mycolactone-dependent structural changes in collagen upon infection with M. ulcerans, associated with the reduction of dermal collagen content, which is in line with our proteomic finding of mycolactone-induced down-regulation of several collagen biosynthesis enzymes. Our results unveil the mechanisms of mycolactone-induced molecular cytopathogenesis on exposed host cells, with the toxin compromising cell structure and homeostasis by inducing cytoskeleton alterations, as well as disrupting tissue structure, by impairing the extracellular matrix biosynthesis.

  4. Proteomic analysis of the action of the Mycobacterium ulcerans toxin mycolactone: targeting host cells cytoskeleton and collagen.

    Science.gov (United States)

    Gama, José B; Ohlmeier, Steffen; Martins, Teresa G; Fraga, Alexandra G; Sampaio-Marques, Belém; Carvalho, Maria A; Proença, Fernanda; Silva, Manuel T; Pedrosa, Jorge; Ludovico, Paula

    2014-08-01

    Buruli ulcer (BU) is a neglected tropical disease caused by Mycobacterium ulcerans. The tissue damage characteristic of BU lesions is known to be driven by the secretion of the potent lipidic exotoxin mycolactone. However, the molecular action of mycolactone on host cell biology mediating cytopathogenesis is not fully understood. Here we applied two-dimensional electrophoresis (2-DE) to identify the mechanisms of mycolactone's cellular action in the L929 mouse fibroblast proteome. This revealed 20 changed spots corresponding to 18 proteins which were clustered mainly into cytoskeleton-related proteins (Dync1i2, Cfl1, Crmp2, Actg1, Stmn1) and collagen biosynthesis enzymes (Plod1, Plod3, P4ha1). In line with cytoskeleton conformational disarrangements that are observed by immunofluorescence, we found several regulators and constituents of both actin- and tubulin-cytoskeleton affected upon exposure to the toxin, providing a novel molecular basis for the effect of mycolactone. Consistent with these cytoskeleton-related alterations, accumulation of autophagosomes as well as an increased protein ubiquitination were observed in mycolactone-treated cells. In vivo analyses in a BU mouse model revealed mycolactone-dependent structural changes in collagen upon infection with M. ulcerans, associated with the reduction of dermal collagen content, which is in line with our proteomic finding of mycolactone-induced down-regulation of several collagen biosynthesis enzymes. Our results unveil the mechanisms of mycolactone-induced molecular cytopathogenesis on exposed host cells, with the toxin compromising cell structure and homeostasis by inducing cytoskeleton alterations, as well as disrupting tissue structure, by impairing the extracellular matrix biosynthesis. PMID:25101965

  5. Remodeling of the Host Cell Plasma Membrane by HIV-1 Nef and Vpu: A Strategy to Ensure Viral Fitness and Persistence

    Directory of Open Access Journals (Sweden)

    Scott M. Sugden

    2016-03-01

    Full Text Available The plasma membrane protects the cell from its surroundings and regulates cellular communication, homing, and metabolism. Not surprisingly, the composition of this membrane is highly controlled through the vesicular trafficking of proteins to and from the cell surface. As intracellular pathogens, most viruses exploit the host plasma membrane to promote viral replication while avoiding immune detection. This is particularly true for the enveloped human immunodeficiency virus (HIV, which assembles and obtains its lipid shell directly at the plasma membrane. HIV-1 encodes two proteins, negative factor (Nef and viral protein U (Vpu, which function primarily by altering the quantity and localization of cell surface molecules to increase virus fitness despite host antiviral immune responses. These proteins are expressed at different stages in the HIV-1 life cycle and employ a variety of mechanisms to target both unique and redundant surface proteins, including the viral receptor CD4, host restriction factors, immunoreceptors, homing molecules, tetraspanins and membrane transporters. In this review, we discuss recent progress in the study of the Nef and Vpu targeting of host membrane proteins with an emphasis on how remodeling of the cell membrane allows HIV-1 to avoid host antiviral immune responses leading to the establishment of systemic and persistent infection.

  6. Altered epigenetic regulation of homeobox genes in human oral squamous cell carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Marcinkiewicz, Katarzyna M.; Gudas, Lorraine J., E-mail: ljgudas@med.cornell.edu

    2014-01-01

    To gain insight into oral squamous cell carcinogenesis, we performed deep sequencing (RNAseq) of non-tumorigenic human OKF6-TERT1R and tumorigenic SCC-9 cells. Numerous homeobox genes are differentially expressed between OKF6-TERT1R and SCC-9 cells. Data from Oncomine, a cancer microarray database, also show that homeobox (HOX) genes are dysregulated in oral SCC patients. The activity of Polycomb repressive complexes (PRC), which causes epigenetic modifications, and retinoic acid (RA) signaling can control HOX gene transcription. HOXB7, HOXC10, HOXC13, and HOXD8 transcripts are higher in SCC-9 than in OKF6-TERT1R cells; using ChIP (chromatin immunoprecipitation) we detected PRC2 protein SUZ12 and the epigenetic H3K27me3 mark on histone H3 at these genes in OKF6-TERT1R, but not in SCC-9 cells. In contrast, IRX1, IRX4, SIX2 and TSHZ3 transcripts are lower in SCC-9 than in OKF6-TERT1R cells. We detected SUZ12 and the H3K27me3 mark at these genes in SCC-9, but not in OKF6-TERT1R cells. SUZ12 depletion increased HOXB7, HOXC10, HOXC13, and HOXD8 transcript levels and decreased the proliferation of OKF6-TERT1R cells. Transcriptional responses to RA are attenuated in SCC-9 versus OKF6-TERT1R cells. SUZ12 and H3K27me3 levels were not altered by RA at these HOX genes in SCC-9 and OKF6-TERT1R cells. We conclude that altered activity of PRC2 is associated with dysregulation of homeobox gene expression in human SCC cells, and that this dysregulation potentially plays a role in the neoplastic transformation of oral keratinocytes. - Highlights: • RNAseq elucidates differences between non-tumorigenic and tumorigenic oral keratinocytes. • Changes in HOX mRNA in SCC-9 vs. OKF6-TERT1R cells are a result of altered epigenetic regulation. • RNAseq shows that retinoic acid (RA) influences gene expression in both OKF6-TERT1R and SCC-9 cells.

  7. Memory CD4+ T cells do not induce graft-versus-host disease.

    Science.gov (United States)

    Anderson, Britt E; McNiff, Jennifer; Yan, Jun; Doyle, Hester; Mamula, Mark; Shlomchik, Mark J; Shlomchik, Warren D

    2003-07-01

    Graft-versus-host disease (GVHD) remains a major cause of morbidity and mortality in allogeneic stem cell transplantation (alloSCT). Donor T cells that accompany stem cell grafts cause GVHD by attacking recipient tissues; therefore, all patients receive GVHD prophylaxis by depletion of T cells from the allograft or through immunosuppressant drugs. In addition to providing a graft-versus-leukemia effect, donor T cells are critical for reconstituting T cell-mediated immunity. Ideally, immunity to infectious agents would be transferred from donor to host without GVHD. Most donors have been exposed to common pathogens and have an increased precursor frequency of memory T cells against pathogenic antigens. We therefore asked whether memory CD62L-CD44+ CD4+ T cells would induce less GVHD than unfractionated or naive CD4+ T cells. Strikingly, we found that memory CD4 cells induced neither clinical nor histologic GVHD. This effect was not due to the increased number of CD4+CD25+ regulatory T cells found in the CD62L-CD44+ fraction because memory T cells depletion of these cells did not cause GVHD. Memory CD4 cells engrafted and responded to antigen both in vivo and in vitro. If these murine results are applicable to human alloSCT, selective administration of memory T cells could greatly improve post-transplant immune reconstitution.

  8. Reduction of fatal graft-versus-host disease by 3H--thymidine suicide of donor cells cultured with host cells

    International Nuclear Information System (INIS)

    The effect of the tritiated thymidine (3H-TdR) suicide technique on the ability of donor cells to induce fatal graft-versus-host disease (GVHD) was studied. C57BL/6 (H-2/sup b/) spleen cells were stimulated in vitro with irradiated BALB/c (H-2/sup d/) Moloney lymphoma cells in mixed culture and 3H-TdR of high-specific activity added to eliminate proliferating cells. The ability of such cells to induce fatal GVHD was assayed by injecting them i.v. into adult BALB/c mice immunosuppressed with cyclophosphamide (180 mg/kg). These cells induced fatal GVHD in fewer mice (52 percent) than did C57BL/6 cells cultured with BALB/c lymphoma cells but without 3H-TdR (87 percent) and C57BL/6 cells cultured with irradiated C57BL/6 cells with (95 percent) or without 3H-TdR (86 percent). Thus, the 3H-TdR suicide technique greatly diminished the ability of cells to induce lethal GVHD

  9. Retinal Targets ALDH Positive Cancer Stem Cell and Alters the Phenotype of Highly Metastatic Osteosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Xiaodong Mu

    2015-01-01

    Full Text Available Aldehyde dehydrogenase (ALDH is a cancer stem cell marker. Retinoic acid has antitumor properties, including the induction of apoptosis and inhibition of proliferation. Retinal, the precursor of retinoic acid, can be oxidized to retinoic acid by dehydrogenases, including ALDH. We hypothesized that retinal could potentially be transformed to retinoic acid with higher efficiency by cancer stem cells, due to the higher ALDH activity. We previously observed that ALDH activity is greater in highly metastatic K7M2 osteosarcoma (OS cells than in nonmetastatic K12 OS cells. We also demonstrated that ALDH activity correlates with clinical metastases in bone sarcoma patients, suggesting that ALDH may be a therapeutic target specific to cells with high metastatic potential. Our current results demonstrated that retinal preferentially affected the phenotypes of ALDH-high K7M2 cells in contrast to ALDH-low K12 cells, which could be mediated by the more efficient transformation of retinal to retinoic acid by ALDH in K7M2 cells. Retinal treatment of highly metastatic K7M2 cells decreased their proliferation, invasion capacity, and resistance to oxidative stress. Retinal altered the expression of metastasis-related genes. These observations indicate that retinal may be used to specifically target metastatic cancer stem cells in OS.

  10. Dynamic Behavior and Function of Foxp3+ Regulatory T Cells in Tumor Bearing Host

    Institute of Scientific and Technical Information of China (English)

    F. Xiao-Feng Qin

    2009-01-01

    Regulatory T cells (Tregs) expressing forkhead/winged-helix transcription factor Foxp3 represent a distinct lineage of lymphocytes which play a central role in protecting the host from autoimmune diseases. However, Tregs also pose a major problem to anti-tumor immunity. Growing body of evidence from both laboratory and clinical investigations has demonstrated that expansion and accumulation of these immunosuppressive cells correlates with advanced tumor growth and predicts poor disease prognosis. How tumor development subverts normal self-tolerance function of Tregs thereby thwarts host anti-tumor immunity remains elusive. This review will discuss our current knowledge in understanding the dynamics and plasticity of Foxp3+ Treg activation and induction in tumor bearing hosts and their interaction with various antigen presenting cells (APCs) in tumor microenvironment leading to the establishment of active local and systemic immune suppression.

  11. Molecular analysis of early host cell infection by Trypanosoma cruzi

    OpenAIRE

    Villalta, Fernando; Madison, M. Nia; Kleshchenko, Yuliya Y.; Nde, Pius N.; Lima, Maria F.

    2008-01-01

    Trypanosoma cruzi, the causative agent of Chagas heart disease, infects heart and other cells leading to cardiac arrest frequently followed by death (1). The disease affects millions of individuals in the Americas and is posing health problems because of blood transmission in the US due to large Latin American immigration (2–3). Since the current drugs present serious side effects and do not cure the chronic infection (4), it is critically important to understand the early process of cellular...

  12. RNAi screen reveals an Abl kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization.

    Directory of Open Access Journals (Sweden)

    Julia F Pielage

    2008-03-01

    Full Text Available Internalization of the pathogenic bacterium Pseudomonas aeruginosa by non-phagocytic cells is promoted by rearrangements of the actin cytoskeleton, but the host pathways usurped by this bacterium are not clearly understood. We used RNAi-mediated gene inactivation of approximately 80 genes known to regulate the actin cytoskeleton in Drosophila S2 cells to identify host molecules essential for entry of P. aeruginosa. This work revealed Abl tyrosine kinase, the adaptor protein Crk, the small GTPases Rac1 and Cdc42, and p21-activated kinase as components of a host signaling pathway that leads to internalization of P. aeruginosa. Using a variety of complementary approaches, we validated the role of this pathway in mammalian cells. Remarkably, ExoS and ExoT, type III secreted toxins of P. aeruginosa, target this pathway by interfering with GTPase function and, in the case of ExoT, by abrogating P. aeruginosa-induced Abl-dependent Crk phosphorylation. Altogether, this work reveals that P. aeruginosa utilizes the Abl pathway for entering host cells and reveals unexpected complexity by which the P. aeruginosa type III secretion system modulates this internalization pathway. Our results furthermore demonstrate the applicability of using RNAi screens to identify host signaling cascades usurped by microbial pathogens that may be potential targets for novel therapies directed against treatment of antibiotic-resistant infections.

  13. Alterations in regulatory T-cells: rediscovered pathways in immunotoxicology

    OpenAIRE

    Corsini, E; Oukka, M; Pieters, R; Kerkvliet, N.I.; Ponce, R.; Germolec, D R

    2011-01-01

    In addition to the effector T-cells subsets, T-cells can also differentiate into cells that play a suppressive or regulatory role in adaptive immune responses. The cell types currently identified as regulatory T-cells (Tregs) include natural or thymic-derived Tregs, T-cells which express Foxp3+CD25+CD4+ and can suppress immune responses to autoreactive T-cells, as well as inducible Tregs, that are generated from naïve T-cells in the periphery after interaction with antigens presented by dendr...

  14. Cell surface appearance of unexpected host MHC determinants on thymocytes from radiation bone marrow chimeras

    International Nuclear Information System (INIS)

    The phenotypic appearance of cell surface antigens on murine thymocytes from long-term radiation bone marrow chimeras was analyzed using indirect immunofluorescence and flow microfluorometry. Cells maturing in the thymi of these mice were typed for MHC (Kk, I-Ak, H-2b, Kb, and Ib) and non-MHC (Lty 1, Ly 9, and TL) determinants. All cells were of donor origin as determined by non-MHC (Ly) phenotype in P1 leads to P2, P1 x P2 leads to P1, and P1 leads to P2 radiation chimeras. In contrast, the MHC phenotypes of these thymocytes were markedly affected by the host environment. Specifically, H-2 and I-A determinants of both parental phenotypes were detected on thymocytes from P1 leads to P1 x P2 chimeras; I-A determinants of host phenotype were present, whereas I-A determinants of donor phenotype were reduced on thymocytes from P1 x P2 leads to P1 chimeras; and thymocytes from P1 leads to P2 chimeras possessed H-2 and I-A determinants of host phenotype but showed reduction of donor I-A phenotype determinants. The appearance of host cell surface H-2 and I-A determinants on thymocytes from chimeras closely parallels the functional recognition of MHC determinants by T cells from chimeric mice and thus may be significantly related to the development of the self-recognition repertoire by maturing T cells

  15. Modulation of host cell signaling pathways as a therapeutic approach in periodontal disease

    Directory of Open Access Journals (Sweden)

    João Antonio Chaves de Souza

    2012-04-01

    Full Text Available Recently, new treatment approaches have been developed to target the host component of periodontal disease. This review aims at providing updated information on host-modulating therapies, focusing on treatment strategies for inhibiting signal transduction pathways involved in inflammation. Pharmacological inhibitors of MAPK, NFκB and JAK/STAT pathways are being developed to manage rheumatoid arthritis, periodontal disease and other inflammatory diseases. Through these agents, inflammatory mediators can be inhibited at cell signaling level, interfering on transcription factors activation and inflammatory gene expression. Although these drugs offer great potential to modulate host response, their main limitations are lack of specificity and developments of side effects. After overcoming these limitations, adjunctive host modulating drugs will provide new therapeutic strategies for periodontal treatment.

  16. Host and habitat filtering in seedling root-associated fungal communities: taxonomic and functional diversity are altered in 'novel' soils.

    Science.gov (United States)

    Pickles, Brian J; Gorzelak, Monika A; Green, D Scott; Egger, Keith N; Massicotte, Hugues B

    2015-10-01

    Climatic and land use changes have significant consequences for the distribution of tree species, both through natural dispersal processes and following management prescriptions. Responses to these changes will be expressed most strongly in seedlings near current species range boundaries. In northern temperate forest ecosystems, where changes are already being observed, ectomycorrhizal fungi contribute significantly to successful tree establishment. We hypothesised that communities of fungal symbionts might therefore play a role in facilitating, or limiting, host seedling range expansion. To test this hypothesis, ectomycorrhizal communities of interior Douglas-fir and interior lodgepole pine seedlings were analysed in a common greenhouse environment following growth in five soils collected along an ecosystem gradient. Currently, Douglas-fir's natural distribution encompasses three of the five soils, whereas lodgepole pine's extends much further north. Host filtering was evident amongst the 29 fungal species encountered: 7 were shared, 9 exclusive to Douglas-fir and 13 exclusive to lodgepole pine. Seedlings of both host species formed symbioses with each soil fungal community, thus Douglas-fir did so even where those soils came from outside its current distribution. However, these latter communities displayed significant taxonomic and functional differences to those found within the host distribution, indicative of habitat filtering. In contrast, lodgepole pine fungal communities displayed high functional similarity across the soil gradient. Taxonomic and/or functional shifts in Douglas-fir fungal communities may prove ecologically significant during the predicted northward migration of this species; especially in combination with changes in climate and management operations, such as seed transfer across geographical regions for forestry purposes. PMID:25694036

  17. Depolarization Alters Phenotype, Maintains Plasticity of Predifferentiated Mesenchymal Stem Cells

    OpenAIRE

    Sundelacruz, Sarah; Levin, Michael; Kaplan, David L

    2013-01-01

    Although adult stem cell transplantation has been implemented as a therapy for tissue repair, it is limited by the availability of functional adult stem cells. A potential approach to generate stem and progenitor cells may be to modulate the differentiated status of somatic cells. Therefore, there is a need for a better understanding of how the differentiated phenotype of mature cells is regulated. We hypothesize that bioelectric signaling plays an important role in the maintenance of the dif...

  18. Secretion of Cpn0796 from Chlamydia pneumoniae into the host cell cytoplasm by an autotransporter mechanism

    DEFF Research Database (Denmark)

    Vandahl, Brian B S; Stensballe, Allan; Roepstorff, Peter;

    2005-01-01

    infected cells, whereas only the 65 kDa full-length Cpn0796 could be detected in purified Chlamydia. Confocal immunofluorescence microscopy demonstrated that Cpn0796 was localized in the Chlamydia membrane in young inclusions. However, at 36 h post infection and later Cpn0796 was detected in the cytoplasm...... of C. pneumoniae infected HEp-2 and BHK cells. Furthermore, Cpn0796 was detected in the cytoplasm of infected cells in the lungs of C. pneumoniae infected C57Bl mice. When cleavage was inhibited, Cpn0796 was retained in the chlamydiae. We propose that Cpn0796 is an autotransporter the N-terminal of...... which is translocated to the host cell cytoplasm. This is the first example of secretion of a Chlamydia autotransporter passenger domain into the host cell cytoplasm. Cpn0796 is specific for C. pneumoniae, where five homologous proteins are encoded by clustered genes. None of these five proteins were...

  19. Mg2Si As Li-Intercalation Host For Li Cells

    Science.gov (United States)

    Huang, Chen-Kuo; Surampudi, Subbarao; Attia, Alan; Halpert, Gerald

    1993-01-01

    Compound Mg2Si shows promise as lithium-intercalation host for ambient-temperature rechargeable lithium electrochemical cells. As anode reactant material, LiXMg2Si chemically stable in presence of organic electrolyte used in such cells and stores large amounts of lithium. Intercalation reactions highly reversible at room temperature. Also retains sufficient mechanical strength during charge/discharge cycling. Lithium cells containing LixMg2Si anodes prove useful in spacecraft, military, communications, automotive, and other applications in which high energy-storage densities of lithium cells in general and rechargeability of cells needed.

  20. Alterations in Cell Signaling Pathways in Breast Cancer Cells after Environmental Exposure

    Energy Technology Data Exchange (ETDEWEB)

    Kulp, K; McCutcheon-Maloney, S M; Bennett, L M

    2003-02-01

    Recent human epidemiological studies suggest that up to 75% of human cancers can be attributed to environmental exposures. Understanding the biologic impact of being exposed to a lifetime of complex environmental mixtures that may not be fully characterized is currently a major challenge. Functional endpoints may be used to assess the gross health consequences of complex mixture exposures from groundwater contamination, superfund sites, biologic releases, or nutritional sources. Such endpoints include the stimulation of cell growth or the induction of a response in an animal model. An environmental exposure that upsets normal cell growth regulation may have important ramifications for cancer development. Stimulating cell growth may alter an individual's cancer risk by changing the expression of genes and proteins that have a role in growth regulatory pathways within cells. Modulating the regulation of these genes and their products may contribute to the initiation, promotion or progression of disease in response to environmental exposure. We are investigating diet-related compounds that induce cell proliferation in breast cancer cell lines. These compounds, PhIP, Flor-Essence{reg_sign} and Essiac{reg_sign}, may be part of an everyday diet. PhIP is a naturally occurring mutagen that is formed in well-cooked muscle meats. PhIP consistently causes dose-dependent breast tumor formation in rats and consumption of well-done meat has been linked to increased risk of breast cancer in women. Flor-Essence{reg_sign} and Essiac{reg_sign} herbal tonics are complementary and alternative medicines used by women who have been diagnosed with breast cancer as an alternative therapy for disease treatment and prevention. The long-term goal of this work is to identify those cellular pathways that are altered by a chemical or biologic environmental exposure and understand how those changes correlate with and or predict changes in human health risk. This project addressed this goal

  1. Adaptation of HIV-1 Depends on the Host-Cell Environment

    Science.gov (United States)

    van Opijnen, Tim; de Ronde, Anthony; Boerlijst, Maarten C.; Berkhout, Ben

    2007-01-01

    Many viruses have the ability to rapidly develop resistance against antiviral drugs and escape from the host immune system. To which extent the host environment affects this adaptive potential of viruses is largely unknown. Here we show that for HIV-1, the host-cell environment is key to the adaptive potential of the virus. We performed a large-scale selection experiment with two HIV-1 strains in two different T-cell lines (MT4 and C8166). Over 110 days of culture, both virus strains adapted rapidly to the MT4 T-cell line. In contrast, when cultured on the C8166 T-cell line, the same strains did not show any increase in fitness. By sequence analyses and infections with viruses expressing either yellow or cyan fluorescent protein, we were able to show that the absence of adaptation was linked to a lower recombination rate in the C8166 T-cell line. Our findings suggest that if we can manipulate the host-cellular factors that mediate viral evolution, we may be able to significantly retard viral adaptability. PMID:17342205

  2. Altering β-cell number through stable alteration of miR-21 and miR-34a expression

    DEFF Research Database (Denmark)

    Backe, Marie Balslev; Novotny, Guy Wayne; Christensen, Dan Ploug;

    2014-01-01

    RNAs, miR-21 and miR-34a, may be involved in mediating cytokine-induced β-cell dysfunction. Therefore, manipulation of miR-21 and miR-34a levels may potentially be beneficial to β cells. To study the effect of long-term alterations of miR-21 or miR-34a levels upon net β-cell number, we stably overexpressed...... miR-21 and knocked down miR-34a, and investigated essential cellular processes. Materials and Methods: miRNA expression was manipulated using Lentiviral transduction of the β-cell line INS-1. Stable cell lines were generated, and cell death, NO synthesis, proliferation, and total cell number were...... monitored in the absence or presence of cytokines. Results: Overexpression of miR-21 decreased net β-cell number in the absence of cytokines, and increased apoptosis and NO synthesis in the absence and presence of cytokines. Proliferation was increased upon miR-21 overexpression. Knockdown of miR-34a...

  3. Trichomonas vaginalis and Tritrichomonas foetus: interaction with fibroblasts and muscle cells - new insights into parasite-mediated host cell cytotoxicity

    Directory of Open Access Journals (Sweden)

    Ricardo Chaves Vilela

    2012-09-01

    Full Text Available Trichomonas vaginalis and Tritrichomonas foetus are parasitic, flagellated protists that inhabit the urogenital tract of humans and bovines, respectively. T. vaginalis causes the most prevalent non-viral sexually transmitted disease worldwide and has been associated with an increased risk for human immunodeficiency virus-1 infection in humans. Infections by T. foetus cause significant losses to the beef industry worldwide due to infertility and spontaneous abortion in cows. Several studies have shown a close association between trichomonads and the epithelium of the urogenital tract. However, little is known concerning the interaction of trichomonads with cells from deeper tissues, such as fibroblasts and muscle cells. Published parasite-host cell interaction studies have reported contradictory results regarding the ability of T. foetus and T. vaginalis to interact with and damage cells of different tissues. In this study, parasite-host cell interactions were examined by culturing primary human fibroblasts obtained from abdominal biopsies performed during plastic surgeries with trichomonads. In addition, mouse 3T3 fibroblasts, primary chick embryo myogenic cells and L6 muscle cells were also used as models of target cells. The parasite-host cell cultures were processed for scanning and transmission electron microscopy and were tested for cell viability and cell death. JC-1 staining, which measures mitochondrial membrane potential, was used to determine whether the parasites induced target cell damage. Terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling staining was used as an indicator of chromatin damage. The colorimetric crystal violet assay was performed to ana-lyse the cytotoxicity induced by the parasite. The results showed that T. foetus and T. vaginalis adhered to and were cytotoxic to both fibroblasts and muscle cells, indicating that trichomonas infection of the connective and muscle tissues is likely to occur; such

  4. Biophysical studies on calcium and carbohydrate binding to carbohydrate recognition domain of Gal/GalNAc lectin from Entamoeba histolytica: insights into host cell adhesion.

    Science.gov (United States)

    Yadav, Rupali; Verma, Kuldeep; Chandra, Mintu; Mukherjee, Madhumita; Datta, Sunando

    2016-09-01

    Entamoeba histolytica, an enteric parasite expresses a Gal/GalNAc-specific lectin that contributes to its virulence by establishing adhesion to host cell. In this study, carbohydrate recognition domain of Hgl (EhCRD) was purified and biophysical studies were conducted to understand the thermodynamic basis of its binding to carbohydrate and Ca(++) Here, we show that carbohydrate recognition domain (CRD) of the lectin binds to calcium through DPN motif. To decipher the role of calcium in carbohydrate binding and host cell adhesion, biophysical and cell-based studies were carried out. We demonstrated that the presence of the cation neither change the affinity of the lectin for carbohydrates nor alters its conformation. Mutation of the calcium-binding motif in EhCRD resulted in complete loss of ability to bind calcium but retained its affinity for carbohydrates. Purified EhCRD significantly diminished adhesion of the amebic trophozoites to Chinese Hamster Ovary (CHO) cells as well as triggered red blood cell agglutination. The calcium-binding defective mutant abrogated amebic adhesion to CHO cells similar to the wild-type protein, but it failed to agglutinate RBCs suggesting a differential role of the cation in these two processes. This study provides the first molecular description of the role of calcium in Gal/GalNAc mediated host cell adhesion.

  5. Biophysical studies on calcium and carbohydrate binding to carbohydrate recognition domain of Gal/GalNAc lectin from Entamoeba histolytica: insights into host cell adhesion.

    Science.gov (United States)

    Yadav, Rupali; Verma, Kuldeep; Chandra, Mintu; Mukherjee, Madhumita; Datta, Sunando

    2016-09-01

    Entamoeba histolytica, an enteric parasite expresses a Gal/GalNAc-specific lectin that contributes to its virulence by establishing adhesion to host cell. In this study, carbohydrate recognition domain of Hgl (EhCRD) was purified and biophysical studies were conducted to understand the thermodynamic basis of its binding to carbohydrate and Ca(++) Here, we show that carbohydrate recognition domain (CRD) of the lectin binds to calcium through DPN motif. To decipher the role of calcium in carbohydrate binding and host cell adhesion, biophysical and cell-based studies were carried out. We demonstrated that the presence of the cation neither change the affinity of the lectin for carbohydrates nor alters its conformation. Mutation of the calcium-binding motif in EhCRD resulted in complete loss of ability to bind calcium but retained its affinity for carbohydrates. Purified EhCRD significantly diminished adhesion of the amebic trophozoites to Chinese Hamster Ovary (CHO) cells as well as triggered red blood cell agglutination. The calcium-binding defective mutant abrogated amebic adhesion to CHO cells similar to the wild-type protein, but it failed to agglutinate RBCs suggesting a differential role of the cation in these two processes. This study provides the first molecular description of the role of calcium in Gal/GalNAc mediated host cell adhesion. PMID:27008865

  6. Alterations in T cell-derived colony-stimulating factors associated with GVH-induced immune deficiency

    International Nuclear Information System (INIS)

    Injection of parental C57BL/10 spleen cells into unirradiated immune-competent (B10 x B10.BR)F1 hosts has been demonstrated to produce a graft-vs.-host-induced immune deficiency in T cell-mediated functions, including mitogen or alloantigen stimulated proliferation or cytotoxic T cell generation. The production of T cell-derived lymphokines affecting hematopoiesis was also altered during GVH. During the first two weeks of GVH, IL-3 and particularly GM-CSF were produced spontaneously; in subsequent weeks, the spontaneous production dropped to normal or subnormal levels. CSF content in concanavalin A-stimulated splenic supernatants was reduced at weeks 1-2, and declined to less than 5% of normal levels by 3-4 weeks of GVH. This decline in CSF content was correlated with a decrease in immune function as assessed by concanavalin A-stimulated IL-2 production and by generation of cytotoxic T lymphocytes. Concurrent with the recovery of immune function during GVH weeks 8-15, mitogen-stimulated production of CSF returned to normal levels. In addition to the decrease in CSF production identified in acute suppressive GVH, CSF content in concanavalin A-stimulated splenic supernatants was also decreased in chronic stimulatory GVH, generated in the strain combination (B6 x B6bm1)F1----(B6bm1 x B6bm12)F1. This decrease in CSF production correlated with a decrease in self-restricted T helper cell function. Finally, a decrease in both immune function and CSF production capacity was observed in the acute GVH following allogeneic (minor histocompatibility loci) bone marrow transplantation into irradiated hosts

  7. Heat-induced alterations in the cell nucleus

    International Nuclear Information System (INIS)

    Hyperthermia may kill eukaryotic cells and may also enhance the radiosensitivity of those cells that survive the heat treatment. Clinically, the possible use of hyperthermia as an adjuvant in the radiotherapeutic treatment of cancer needs the understanding of mechanisms that underlay heat-induced cell death and radiosensitization. By in vitro heating of established human (HeLaS3) and rodent (Ehrlich Ascites Tumor and LM fibroblast) cell lines, both killing and radiosensitization were investigated. (author). 1067 refs.; 76 figs.; 19 tabs

  8. Alemtuzumab treatment alters circulating innate immune cells in multiple sclerosis

    Science.gov (United States)

    Ahmetspahic, Diana; Ruck, Tobias; Schulte-Mecklenbeck, Andreas; Schwarte, Kathrin; Jörgens, Silke; Scheu, Stefanie; Windhagen, Susanne; Graefe, Bettina; Melzer, Nico; Klotz, Luisa; Arolt, Volker; Wiendl, Heinz; Meuth, Sven G.

    2016-01-01

    Objective: To characterize changes in myeloid and lymphoid innate immune cells in patients with relapsing-remitting multiple sclerosis (MS) during a 6-month follow-up after alemtuzumab treatment. Methods: Circulating innate immune cells including myeloid cells and innate lymphoid cells (ILCs) were analyzed before and 6 and 12 months after onset of alemtuzumab treatment. Furthermore, a potential effect on granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)–23 production by myeloid cells and natural killer (NK) cell cytolytic activity was determined. Results: In comparison to CD4+ T lymphocytes, myeloid and lymphoid innate cell subsets of patients with MS expressed significantly lower amounts of CD52 on their cell surface. Six months after CD52 depletion, numbers of circulating plasmacytoid dendritic cells (DCs) and conventional DCs were reduced compared to baseline. GM-CSF and IL-23 production in DCs remained unchanged. Within the ILC compartment, the subset of CD56bright NK cells specifically expanded under alemtuzumab treatment, but their cytolytic activity did not change. Conclusions: Our findings demonstrate that 6 months after alemtuzumab treatment, specific DC subsets are reduced, while CD56bright NK cells expanded in patients with MS. Thus, alemtuzumab specifically restricts the DC compartment and expands the CD56bright NK cell subset with potential immunoregulatory properties in MS. We suggest that remodeling of the innate immune compartment may promote long-term efficacy of alemtuzumab and preserve immunocompetence in patients with MS. PMID:27766281

  9. Anaplasma phagocytophilum Manipulates Host Cell Apoptosis by Different Mechanisms to Establish Infection

    Directory of Open Access Journals (Sweden)

    Pilar Alberdi

    2016-07-01

    Full Text Available Anaplasma phagocytophilum is an emerging zoonotic pathogen that causes human and animal granulocytic anaplasmosis and tick-borne fever of ruminants. This obligate intracellular bacterium evolved to use common strategies to establish infection in both vertebrate hosts and tick vectors. Herein, we discuss the different strategies used by the pathogen to modulate cell apoptosis and establish infection in host cells. In vertebrate neutrophils and human promyelocytic cells HL-60, both pro-apoptotic and anti-apoptotic factors have been reported. Tissue-specific differences in tick response to infection and differential regulation of apoptosis pathways have been observed in adult female midguts and salivary glands in response to infection with A. phagocytophilum. In tick midguts, pathogen inhibits apoptosis through the Janus kinase/signal transducers and activators of transcription (JAK/STAT pathway, while in salivary glands, the intrinsic apoptosis pathways is inhibited but tick cells respond with the activation of the extrinsic apoptosis pathway. In Ixodes scapularis ISE6 cells, bacterial infection down-regulates mitochondrial porin and manipulates protein processing in the endoplasmic reticulum and cell glucose metabolism to inhibit apoptosis and facilitate infection, whereas in IRE/CTVM20 tick cells, inhibition of apoptosis appears to be regulated by lower caspase levels. These results suggest that A. phagocytophilum uses different mechanisms to inhibit apoptosis for infection of both vertebrate and invertebrate hosts.

  10. Interrelations between the Parasitophorous Vacuole of Toxoplasma gondii and Host Cell Organelles

    Science.gov (United States)

    Cardoso Magno, Rodrigo; Cobra Straker, Lorian; de Souza, Wanderley; Attias, Marcia

    2005-04-01

    Toxoplasma gondii, the causative agent of toxoplasmosis, is capable of actively penetrating and multiplying in any nucleated cell of warm-blooded animals. Its survival strategies include escape from fusion of the parasitophorous vacuole with host cell lysosomes and rearrangement of host cell organelles in relation to the parasitophorous vacuole. In this article we report the rearrangement of host cell organelles and elements of the cytoskeleton of LLCMK2 cells, a lineage derived from green monkey kidney epithelial cells, in response to infection by T. gondii tachyzoites. Transmission electron microscopy made on flat embedded monolayers cut horizontally to the apical side of the cells or field emission scanning electron microscopy of monolayers scraped with scotch tape before sputtering showed that association of mitochondria to the vacuole is much less frequent than previously described. On the other hand, all parasitophorous vacuoles were surrounded by elements of the endoplasmic reticulum. These data were complemented by observations by laser scanning microscopy using fluorescent probes from mitochondria and endoplasmic reticulum and reinforced by three-dimensional reconstruction from serial sections observed by transmission electron microscopy and labeling of mitochondria and endoplasmic reticulum by fluorescent probes.

  11. Industrial production of clotting factors: Challenges of expression, and choice of host cells.

    Science.gov (United States)

    Kumar, Sampath R

    2015-07-01

    The development of recombinant forms of blood coagulation factors as safer alternatives to plasma derived factors marked a major advance in the treatment of common coagulation disorders. These are complex proteins, mostly enzymes or co-enzymes, involving multiple post-translational modifications, and therefore are difficult to express. This article reviews the nature of the expression challenges for the industrial production of these factors, vis-à-vis the translational and post-translational bottlenecks, as well as the choice of host cell lines for high-fidelity production. For achieving high productivities of vitamin K dependent proteins, which include factors II (prothrombin), VII, IX and X, and protein C, host cell limitation of γ-glutamyl carboxylation is a major bottleneck. Despite progress in addressing this, involvement of yet unidentified protein(s) impedes a complete cell engineering solution. Human factor VIII expresses at very low levels due to limitations at several steps in the protein secretion pathway. Protein and cell engineering, vector improvement and alternate host cells promise improvement in the productivity. Production of Von Willebrand factor is constrained by its large size, complex structure, and the need for extensive glycosylation and disulfide-bonded oligomerization. All the licensed therapeutic factors are produced in CHO, BHK or HEK293 cells. While HEK293 is a recent adoption, BHK cells appear to be disfavored. PMID:26099845

  12. A novel mechanism of bacterial toxin transfer within host blood cell-derived microvesicles.

    Directory of Open Access Journals (Sweden)

    Anne-lie Ståhl

    2015-02-01

    Full Text Available Shiga toxin (Stx is the main virulence factor of enterohemorrhagic Escherichia coli, which are non-invasive strains that can lead to hemolytic uremic syndrome (HUS, associated with renal failure and death. Although bacteremia does not occur, bacterial virulence factors gain access to the circulation and are thereafter presumed to cause target organ damage. Stx was previously shown to circulate bound to blood cells but the mechanism by which it would potentially transfer to target organ cells has not been elucidated. Here we show that blood cell-derived microvesicles, shed during HUS, contain Stx and are found within patient renal cortical cells. The finding was reproduced in mice infected with Stx-producing Escherichia coli exhibiting Stx-containing blood cell-derived microvesicles in the circulation that reached the kidney where they were transferred into glomerular and peritubular capillary endothelial cells and further through their basement membranes followed by podocytes and tubular epithelial cells, respectively. In vitro studies demonstrated that blood cell-derived microvesicles containing Stx undergo endocytosis in glomerular endothelial cells leading to cell death secondary to inhibited protein synthesis. This study demonstrates a novel virulence mechanism whereby bacterial toxin is transferred within host blood cell-derived microvesicles in which it may evade the host immune system.

  13. Where in the Cell Are You? Probing HIV-1 Host Interactions through Advanced Imaging Techniques

    Science.gov (United States)

    Dirk, Brennan S.; Van Nynatten, Logan R.; Dikeakos, Jimmy D.

    2016-01-01

    Viruses must continuously evolve to hijack the host cell machinery in order to successfully replicate and orchestrate key interactions that support their persistence. The type-1 human immunodeficiency virus (HIV-1) is a prime example of viral persistence within the host, having plagued the human population for decades. In recent years, advances in cellular imaging and molecular biology have aided the elucidation of key steps mediating the HIV-1 lifecycle and viral pathogenesis. Super-resolution imaging techniques such as stimulated emission depletion (STED) and photoactivation and localization microscopy (PALM) have been instrumental in studying viral assembly and release through both cell–cell transmission and cell–free viral transmission. Moreover, powerful methods such as Forster resonance energy transfer (FRET) and bimolecular fluorescence complementation (BiFC) have shed light on the protein-protein interactions HIV-1 engages within the host to hijack the cellular machinery. Specific advancements in live cell imaging in combination with the use of multicolor viral particles have become indispensable to unravelling the dynamic nature of these virus-host interactions. In the current review, we outline novel imaging methods that have been used to study the HIV-1 lifecycle and highlight advancements in the cell culture models developed to enhance our understanding of the HIV-1 lifecycle. PMID:27775563

  14. In situ tissue engineering of functional small-diameter blood vessels by host circulating cells only

    NARCIS (Netherlands)

    Talacua, Hanna; Smits, Anthal I P M; Muylaert, Dimitri E P; Van Rijswijk, Jan Willem; Vink, Aryan; Verhaar, Marianne C.; Driessen-Mol, Anita; Van Herwerden, Lex A.; Bouten, Carlijn V C; Kluin, Jolanda; Baaijens, Frank P T

    2015-01-01

    Inflammation is a natural phase of the wound healing response, which can be harnessed for the in situ tissue engineering of small-diameter blood vessels using instructive, bioresorbable synthetic grafts. This process is dependent on colonization of the graft by host circulating cells and subsequent

  15. Intracellular accommodation of rhizobia in legume host cell: the fine-tuning of the endomembrane system

    NARCIS (Netherlands)

    Gavrin, A.Y.

    2015-01-01

    The symbiosis of legumes with rhizobia leads to the formation of root nodules. Rhizobia which are hosted inside specialized infected cells are surrounded by hostderived membranes, forming symbiosomes. Although it is known that symbiosome formation involves proliferation of membranes and changing of

  16. Altered effector function of peripheral cytotoxic cells in COPD

    Directory of Open Access Journals (Sweden)

    Corne Jonathan M

    2009-06-01

    Full Text Available Abstract Background There is mounting evidence that perforin and granzymes are important mediators in the lung destruction seen in COPD. We investigated the characteristics of the three main perforin and granzyme containing peripheral cells, namely CD8+ T lymphocytes, natural killer (NK; CD56+CD3- cells and NKT-like (CD56+CD3+ cells. Methods Peripheral blood mononuclear cells (PBMCs were isolated and cell numbers and intracellular granzyme B and perforin were analysed by flow cytometry. Immunomagnetically selected CD8+ T lymphocytes, NK (CD56+CD3- and NKT-like (CD56+CD3+ cells were used in an LDH release assay to determine cytotoxicity and cytotoxic mechanisms were investigated by blocking perforin and granzyme B with relevant antibodies. Results The proportion of peripheral blood NKT-like (CD56+CD3+ cells in smokers with COPD (COPD subjects was significantly lower (0.6% than in healthy smokers (smokers (2.8%, p +CD3- cells from COPD subjects were significantly less cytotoxic than in smokers (16.8% vs 51.9% specific lysis, p +CD3+ cells (16.7% vs 52.4% specific lysis, p +CD3- and NKT-like (CD56+CD3+ cells from smokers and HNS. Conclusion In this study, we show that the relative numbers of peripheral blood NK (CD56+CD3- and NKT-like (CD56+CD3+ cells in COPD subjects are reduced and that their cytotoxic effector function is defective.

  17. The Paracoccidioides cell wall: past and present layers towards understanding interaction with the host

    Directory of Open Access Journals (Sweden)

    Rosana ePuccia

    2011-12-01

    Full Text Available The cell wall of pathogenic fungi plays import roles in interaction with the host, so that its composition and structure may determine the course of infection. Here we present an overview of the current and past knowledge on the cell wall constituents of Paracoccidioides brasiliensis and P. lutzii. These are temperature-dependent dimorphic fungi that cause paracoccidioidomycosis, a systemic granulomatous and debilitating disease. Focus is given on cell wall carbohydrate and protein contents, their immune-stimulatory features, adhesion properties, drug target characteristics, and morphological phase specificity. We offer a journey towards the future understanding of the dynamic life that takes place in the cell wall and of the changes that it may suffer when living in the human host.

  18. Next-Generation "-omics" Approaches Reveal a Massive Alteration of Host RNA Metabolism during Bacteriophage Infection of Pseudomonas aeruginosa.

    Science.gov (United States)

    Chevallereau, Anne; Blasdel, Bob G; De Smet, Jeroen; Monot, Marc; Zimmermann, Michael; Kogadeeva, Maria; Sauer, Uwe; Jorth, Peter; Whiteley, Marvin; Debarbieux, Laurent; Lavigne, Rob

    2016-07-01

    As interest in the therapeutic and biotechnological potentials of bacteriophages has grown, so has value in understanding their basic biology. However, detailed knowledge of infection cycles has been limited to a small number of model bacteriophages, mostly infecting Escherichia coli. We present here the first analysis coupling data obtained from global next-generation approaches, RNA-Sequencing and metabolomics, to characterize interactions between the virulent bacteriophage PAK_P3 and its host Pseudomonas aeruginosa. We detected a dramatic global depletion of bacterial transcripts coupled with their replacement by viral RNAs over the course of infection, eventually leading to drastic changes in pyrimidine metabolism. This process relies on host machinery hijacking as suggested by the strong up-regulation of one bacterial operon involved in RNA processing. Moreover, we found that RNA-based regulation plays a central role in PAK_P3 lifecycle as antisense transcripts are produced mainly during the early stage of infection and viral small non coding RNAs are massively expressed at the end of infection. This work highlights the prominent role of RNA metabolism in the infection strategy of a bacteriophage belonging to a new characterized sub-family of viruses with promising therapeutic potential. PMID:27380413

  19. Next-Generation "-omics" Approaches Reveal a Massive Alteration of Host RNA Metabolism during Bacteriophage Infection of Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Anne Chevallereau

    2016-07-01

    Full Text Available As interest in the therapeutic and biotechnological potentials of bacteriophages has grown, so has value in understanding their basic biology. However, detailed knowledge of infection cycles has been limited to a small number of model bacteriophages, mostly infecting Escherichia coli. We present here the first analysis coupling data obtained from global next-generation approaches, RNA-Sequencing and metabolomics, to characterize interactions between the virulent bacteriophage PAK_P3 and its host Pseudomonas aeruginosa. We detected a dramatic global depletion of bacterial transcripts coupled with their replacement by viral RNAs over the course of infection, eventually leading to drastic changes in pyrimidine metabolism. This process relies on host machinery hijacking as suggested by the strong up-regulation of one bacterial operon involved in RNA processing. Moreover, we found that RNA-based regulation plays a central role in PAK_P3 lifecycle as antisense transcripts are produced mainly during the early stage of infection and viral small non coding RNAs are massively expressed at the end of infection. This work highlights the prominent role of RNA metabolism in the infection strategy of a bacteriophage belonging to a new characterized sub-family of viruses with promising therapeutic potential.

  20. Aedes aegypti saliva alters leukocyte recruitment and cytokine signaling by antigen-presenting cells during West Nile virus infection.

    Directory of Open Access Journals (Sweden)

    Bradley S Schneider

    Full Text Available West Nile virus (WNV is transmitted during mosquito bloodfeeding. Consequently, the first vertebrate cells to contact WNV are cells in the skin, followed by those in the draining lymph node. Macrophages and dendritic cells are critical early responders in host defense against WNV infection, not just because of their role in orchestrating the immune response, but also because of their importance as sites of early peripheral viral replication. Antigen-presenting cell (APC signals have a profound effect on host antiviral responses and disease severity. During transmission, WNV is intimately associated with mosquito saliva. Due to the ability of mosquito saliva to affect inflammation and immune responses, and the importance of understanding early events in WNV infection, we investigated whether mosquito saliva alters APC signaling during arbovirus infection, and if alterations in cell recruitment occur when WNV infection is initiated with mosquito saliva. Accordingly, experiments were performed with cultured dendritic cells and macrophages, flow cytometry was used to characterize infiltrating cell types in the skin and lymph nodes during early infection, and real-time RT-PCR was employed to evaluate virus and cytokine levels. Our in vitro results suggest that mosquito saliva significantly decreases the expression of interferon-beta and inducible nitric oxide synthase in macrophages (by as much as 50 and 70%, respectively, whilst transiently enhancing interleukin-10 (IL-10 expression. In vivo results indicate that the predominate effect of mosquito feeding is to significantly reduce the recruitment of T cells, leading the inoculation site of mice exposed to WNV alone to have up to 2.8 fold more t cells as mice infected in the presence of mosquito saliva. These shifts in cell population are associated with significantly elevated IL-10 and WNV (up to 4.0 and 10 fold, respectively in the skin and draining lymph nodes. These results suggest that mosquito

  1. B7-H3 expression in donor T cells and host cells negatively regulates acute graft-versus-host disease lethality.

    Science.gov (United States)

    Veenstra, Rachelle G; Flynn, Ryan; Kreymborg, Katharina; McDonald-Hyman, Cameron; Saha, Asim; Taylor, Patricia A; Osborn, Mark J; Panoskaltsis-Mortari, Angela; Schmitt-Graeff, Annette; Lieberknect, Elisabeth; Murphy, William J; Serody, Jonathan S; Munn, David H; Freeman, Gordon J; Allison, James P; Mak, Tak W; van den Brink, Marcel; Zeiser, Robert; Blazar, Bruce R

    2015-05-21

    Members of the B7 family have been shown to be important for regulating immune responses by providing either positive or negative costimulatory signals. The function of B7-H3 has been controversial. We show that B7-H3 is upregulated in graft-versus-host disease (GVHD) target organs, including the colon, liver, and lung. Infusion of allogeneic donor T cells into B7-H3(-/-) vs wild-type (WT) recipients resulted in increased GVHD lethality associated with increased T-cell proliferation, colonic inflammatory cytokines, and destruction of epithelial barriers. Allogeneic B7-H3(-/-) vs WT donor T cells also had increased T-cell proliferation and GVHD lethality associated with increased proliferation and cytokine secretion in the spleen, intraepithelial lymphocyte inflammatory cytokines, and intestinal permeability. Both resting and activated regulatory T cells (Tregs) lack B7-H3 messenger RNA. Consistent with these data, GVHD was augmented in recipients of B7-H3(-/-) Treg-depleted grafts. In two delayed lymphocyte infusion (DLI) models, T cells lacking B7-H3 are capable of providing graft-versus-leukemia (GVL) effects. We conclude that B7-H3 is responsible for providing a negative costimulatory signal. Our studies provide support for developing and testing new therapies directed toward the B7-H3 pathway, including approaches to augment host B7-H3 early after bone marrow transplantation to prevent GVHD and to develop potent antagonistic antibodies later after transplant to facilitate DLI-mediated GVL without GVHD complications. PMID:25814530

  2. Novel insights into host-fungal pathogen interactions derived from live-cell imaging.

    Science.gov (United States)

    Bain, Judith; Gow, Neil A R; Erwig, Lars-Peter

    2015-03-01

    The theoretical physicist and Nobel laureate Richard Feynman outlined in his 1959 lecture, "There's plenty of room at the bottom", the enormous possibility of producing and visualising things at smaller scales. The advent of advanced scanning and transmission electron microscopy and high-resolution microscopy has begun to open the door to visualise host-pathogen interactions at smaller scales, and spinning disc confocal and two-photon microscopy has improved our ability to study these events in real time in three dimensions. The aim of this review is to illustrate some of the advances in understanding host-fungal interactions that have been made in recent years in particular those relating to the interactions of live fungal pathogens with phagocytes. Dynamic imaging of host-pathogen interactions has recently revealed novel detail and unsuspected mechanistic insights, facilitating the dissection of the phagocytic process into its component parts. Here, we will highlight advances in our knowledge of host-fungal pathogen interactions, including the specific effects of fungal cell viability, cell wall composition and morphogenesis on the phagocytic process and try to define the relative contributions of neutrophils and macrophages to the clearance of fungal pathogens in vitro and the infected host.

  3. Dendritic cell biology in human cytomegalovirus infection and the clinical consequences for host immunity and pathology

    OpenAIRE

    Gredmark-Russ, Sara; Söderberg-Nauclér, Cecilia

    2012-01-01

    Human cytomegalovirus (HCMV), a member of the herpesvirus family, establishes life-long persistence and latency after primary infection and can be reactivated later in life. In immunosuppressed patients, it is an important pathogen that can cause severe disease. HCMV is also thought to play a causative role in inflammatory diseases and cancer. The virus can infect different immune cells, including dendritic cells (DCs) and can take advantage of host immune functions to avoid immune recognitio...

  4. Lymphotoxin organizes contributions to host defense and metabolic illness from innate lymphoid cells

    OpenAIRE

    Upadhyay, Vaibhav; Fu, Yang-Xin

    2013-01-01

    The lymphotoxin (LT)-pathway is a unique constituent branch of the Tumor Necrosis Superfamily (TNFSF). Use of LT is a critical mechanism by which fetal innate lymphoid cells regulate lymphoid organogenesis. Within recent years, adult innate lymphoid cells have been discovered to utilize this same pathway to regulate IL-22 and IL-23 production for host defense. Notably, genetic studies have linked polymorphisms in the genes encoding LTα to several phenotypes contributing to metabolic syndrome....

  5. Ultrastructural characteristics of nurse cell-larva complex of four species of Trichinella in several hosts

    OpenAIRE

    Sacchi L.; Corona S.; Gajadhar A.A.; Pozio E.

    2001-01-01

    The nurse cell-larva complex of nematodes of the genus Trichinella plays an Important role in the survival of the larva in decaying muscles, frequently favouring the transmission of the parasite in extreme environmental conditions. The ultrastructure of the nurse cell-larva complex in muscles from different hosts infected with T. nativa (a walrus and a polar bear), T. spiralis (horses and humans), T. pseudospiralis (a laboratory mouse) and T. papuae (a laboratory mouse) were examined. Analysi...

  6. Patterns of Oligonucleotide Sequences in Viral and Host Cell RNA Identify Mediators of the Host Innate Immune System

    NARCIS (Netherlands)

    Greenbaum, Benjamin D.; Rabadan, Raul; Levine, Arnold J.

    2009-01-01

    The innate immune response provides a first line of defense against pathogens by targeting generic differential features that are present in foreign organisms but not in the host. These innate responses generate selection forces acting both in pathogens and hosts that further determine their co-evol

  7. Serratia marcescens induces apoptotic cell death in host immune cells via a lipopolysaccharide- and flagella-dependent mechanism.

    Science.gov (United States)

    Ishii, Kenichi; Adachi, Tatsuo; Imamura, Katsutoshi; Takano, Shinya; Usui, Kimihito; Suzuki, Kazushi; Hamamoto, Hiroshi; Watanabe, Takeshi; Sekimizu, Kazuhisa

    2012-10-19

    Injection of Serratia marcescens into the blood (hemolymph) of the silkworm, Bombyx mori, induced the activation of c-Jun NH(2)-terminal kinase (JNK), followed by caspase activation and apoptosis of blood cells (hemocytes). This process impaired the innate immune response in which pathogen cell wall components, such as glucan, stimulate hemocytes, leading to the activation of insect cytokine paralytic peptide. S. marcescens induced apoptotic cell death of silkworm hemocytes and mouse peritoneal macrophages in vitro. We searched for S. marcescens transposon mutants with attenuated ability to induce apoptosis of silkworm hemocytes. Among the genes identified, disruption mutants of wecA (a gene involved in lipopolysaccharide O-antigen synthesis), and flhD and fliR (essential genes in flagella synthesis) showed reduced motility and impaired induction of mouse macrophage cell death. These findings suggest that S. marcescens induces apoptosis of host immune cells via lipopolysaccharide- and flagella-dependent motility, leading to the suppression of host innate immunity.

  8. Heparanase promotes engraftment and prevents graft versus host disease in stem cell transplantation.

    Directory of Open Access Journals (Sweden)

    Menachem Bitan

    Full Text Available BACKGROUND: Heparanase, endoglycosidase that cleaves heparan sulfate side chains of heparan sulfate proteoglycans, plays important roles in cancer metastasis, angiogenesis and inflammation. DESIGN AND METHODS: Applying a mouse model of bone marrow transplantation and transgenic mice over-expressing heparanase, we evaluated the effect of heparanase on the engraftment process and the development of graft-versus-host disease. RESULTS: Analysis of F1 mice undergoing allogeneic bone marrow transplantation from C57BL/6 mice demonstrated a better and faster engraftment in mice receiving cells from donors that were pretreated with heparanase. Moreover, heparanase treated recipient F1 mice showed only a mild appearance of graft-versus-host disease and died 27 days post transplantation while control mice rapidly developed signs of graft-versus-host disease (i.e., weight loss, hair loss, diarrhea and died after 12 days, indicating a protective effect of heparanase against graft-versus-host disease. Similarly, we applied transgenic mice over-expressing heparanase in most tissues as the recipients of BMT from C57BL/6 mice. Monitoring clinical parameters of graft-versus-host disease, the transgenic mice showed 100% survival on day 40 post transplantation, compared to only 50% survival on day 14, in the control group. In vitro and in vivo studies revealed that heparanase inhibited T cell function and activation through modulation of their cytokine repertoire, indicated by a marked increase in the levels of Interleukin-4, Interleukin-6 and Interleukin-10, and a parallel decrease in Interleukin-12, tumor necrosis factor-alfa and interferon-gamma. Using point mutated inactive enzyme, we found that the shift in cytokine profile was independent of heparanase enzymatic activity. CONCLUSIONS: Our results indicate a significant role of heparanase in bone marrow transplantation biology, facilitating engraftment and suppressing graft-versus-host disease, apparently

  9. Carbofuran alters centrosome and spindle organization, and delays cell division in oocytes and mitotic cells.

    Science.gov (United States)

    Cinar, Ozgur; Semiz, Olcay; Can, Alp

    2015-04-01

    Although many countries banned of its usage, carbofuran (CF) is still one of the most commonly used carbamate derivative insecticides against insects and nematodes in agriculture and household, threatening the human and animal health by contaminating air, water, and food. Our goal was to evaluate the potential toxic effects of CF on mammalian oocytes besides mitotic cells. Caspase-dependent apoptotic pathway was assessed by immunofluorescence and western blot techniques. Alterations in the meiotic spindle formation after CF exposure throughout the in vitro maturation of mice oocyte-cumulus complexes (COCs) were analyzed by using a 3D confocal laser microscope. Maturation efficiency and kinetics were assessed by direct observation of the COCs. Results indicated that the number of TUNEL-positive cells increased in CF-exposed groups, particularly higher doses (>250 µM) in a dose-dependent fashion. The ratio of anticleaved caspase-3 labeled cells in those groups positively correlated with TUNEL-positivity. Western blot analysis confirmed a significant increase in active caspase-3 activity. CF caused a dose-dependent accumulation of oocytes at prometaphase-I (PM-I) of meiosis. Partial loss of spindle microtubules (MTs) was noted, which consequently gave rise to a diamond shape spindle. Aberrant pericentrin foci were noted particularly in PM-I and metaphase-I (M-I) stages. Conclusively, CF (1) induces programmed cell death in a dose-dependent manner, and (2) alters spindle morphology most likely through a mechanism that interacts with MT assembly and/or disorientation of pericentriolar proteins. Overall, data suggest that CF could give rise to aneuploidy or cell death in higher doses, therefore reduce fertilization and implantation rates.

  10. The cytoskeleton adaptor protein ankyrin-1 is upregulated by p53 following DNA damage and alters cell migration.

    Science.gov (United States)

    Hall, A E; Lu, W-T; Godfrey, J D; Antonov, A V; Paicu, C; Moxon, S; Dalmay, T; Wilczynska, A; Muller, P A J; Bushell, M

    2016-01-01

    The integrity of the genome is maintained by a host of surveillance and repair mechanisms that are pivotal for cellular function. The tumour suppressor protein p53 is a major component of the DNA damage response pathway and plays a vital role in the maintenance of cell-cycle checkpoints. Here we show that a microRNA, miR-486, and its host gene ankyrin-1 (ANK1) are induced by p53 following DNA damage. Strikingly, the cytoskeleton adaptor protein ankyrin-1 was induced over 80-fold following DNA damage. ANK1 is upregulated in response to a variety of DNA damage agents in a range of cell types. We demonstrate that miR-486-5p is involved in controlling G1/S transition following DNA damage, whereas the induction of the ankyrin-1 protein alters the structure of the actin cytoskeleton and sustains limited cell migration during DNA damage. Importantly, we found that higher ANK1 expression correlates with decreased survival in cancer patients. Thus, these observations highlight ANK1 as an important effector downstream of the p53 pathway. PMID:27054339

  11. Lower biodiversity of native fish but only marginally altered plankton biomass in tropical lakes hosting introduced piscivorous Cichla cf. ocellaris

    DEFF Research Database (Denmark)

    Menezes, Rosemberg; Attayde, José Luiz; Lacerot, Gissell;

    2012-01-01

    We compared the species richness and abundance of fish, zooplankton and phytoplankton in nine mesotrophic coastal shallow lakes (Northeastern Brazil) with and without the exotic predator cichlid tucunaré or ‘peacock bass’ (Cichla cf. ocellaris). We hypothesized that the introduction of tucunaré...... would lead to decreased abundance and species diversity of native fish assemblages and cause indirect effects on the abundance and species diversity of the existing communities of zooplankton and phytoplankton and on water transparency. Our hypotheses were only partly confirmed. Although fish richness...... and diversity were, in fact, drastically lower in the lakes hosting tucunaré, no significant differences were traced in total fish catch per unit of effort, zooplankton and phytoplankton biomass, plankton diversity or the zooplankton:phytoplankton biomass (TZOO:TPHYTO) ratio. However, zooplankton biomass...

  12. Cell surface glycan alterations in epithelial mesenchymal transition process of Huh7 hepatocellular carcinoma cell.

    Directory of Open Access Journals (Sweden)

    Shan Li

    Full Text Available BACKGROUND AND OBJECTIVE: Due to recurrence and metastasis, the mortality of Hepatocellular carcinoma (HCC is high. It is well known that the epithelial mesenchymal transition (EMT and glycan of cell surface glycoproteins play pivotal roles in tumor metastasis. The goal of this study was to identify HCC metastasis related differential glycan pattern and their enzymatic basis using a HGF induced EMT model. METHODOLOGY: HGF was used to induce HCC EMT model. Lectin microarray was used to detect the expression of cell surface glycan and the difference was validated by lectin blot and fluorescence cell lectin-immunochemistry. The mRNA expression levels of glycotransferases were determined by qRT-PCR. RESULTS: After HGF treatment, the Huh7 cell lost epithelial characteristics and obtained mesenchymal markers. These changes demonstrated that HGF could induce a typical cell model of EMT. Lectin microarray analysis identified a decreased affinity in seven lectins ACL, BPL, JAC, MPL, PHA-E, SNA, and SBA to the glycan of cell surface glycoproteins. This implied that glycan containing T/Tn-antigen, NA2 and bisecting GlcNAc, Siaα2-6Gal/GalNAc, terminal α or βGalNAc structures were reduced. The binding ability of thirteen lectins, AAL, LCA, LTL, ConA, NML, NPL, DBA, HAL, PTL II, WFL, ECL, GSL II and PHA-L to glycan were elevated, and a definite indication that glycan containing terminal αFuc and ± Sia-Le, core fucose, α-man, gal-β(α GalNAc, β1,6 GlcNAc branching and tetraantennary complex oligosaccharides structures were increased. These results were further validated by lectin blot and fluorescence cell lectin-immunochemistry. Furthermore, the mRNA expression level of Mgat3 decreased while that of Mgat5, FucT8 and β3GalT5 increased. Therefore, cell surface glycan alterations in the EMT process may coincide with the expression of glycosyltransferase. CONCLUSIONS: The findings of this study systematically clarify the alterations of cell surface

  13. Transfected parvalbumin alters calcium homeostasis in teratocarcinoma PCC7 cells

    DEFF Research Database (Denmark)

    Müller, B K; Kabos, P; Belhage, B;

    1996-01-01

    transfected. Parvalbumin-transfected and mock-transfected cells were loaded with the calcium indicator fura-2 and were exposed, in the same dish, to different concentrations of the calcium ionophore A23187 or to KCI. The results show that parvalbumin-transfected PCC7 cells had much better calcium buffering...

  14. Effects of altered gravity on the cell cycle, actin cytoskeleton and proteome in Physarum polycephalum

    Science.gov (United States)

    He, Jie; Zhang, Xiaoxian; Gao, Yong; Li, Shuijie; Sun, Yeqing

    Some researchers suggest that the changes of cell cycle under the effect of microgravity may be associated with many serious adverse physiological changes. In the search for underlying mechanisms and possible new countermeasures, we used the slime mold Physarum polycephalum in which all the nuclei traverse the cell cycle in natural synchrony to study the effects of altered gravity on the cell cycle, actin cytoskeleton and proteome. In parallel, the cell cycle was analyzed in Physarum incubated (1) in altered gravity for 20 h, (2) in altered gravity for 40 h, (3) in altered gravity for 80 h, and (4) in ground controls. The cell cycle, the actin cytoskeleton, and proteome in the altered gravity and ground controls were examined. The results indicated that the duration of the G2 phase was lengthened 20 min in high aspect ratio vessel (HARV) for 20 h, and prolonged 2 h in altered gravity either for 40 h or for 80 h, whereas the duration of other phases in the cell cycle was unchanged with respect to the control. The microfilaments in G2 phase had a reduced number of fibers and a unique abnormal morphology in altered gravity for 40 h, whereas the microfilaments in other phases of cell cycle were unchanged when compared to controls. Employing classical two-dimensional electrophoresis (2-DE), we examined the effect of the altered gravity on P. polycephalum proteins. The increase in the duration of G2 phase in altered gravity for 40 h was accompanied by changes in the 2-DE protein profiles, over controls. Out of a total of 200 protein spots investigated in G2 phase, which were reproducible in repeated experiments, 72 protein spots were visually identified as specially expressed, and 11 proteins were up-regulated by 2-fold and 28 proteins were down-regulated by 2-fold over controls. Out of a total of three low-expressed proteins in G2 phase in altered gravity for 40 h, two proteins were unknown proteins, and one protein was spherulin 3b by MALDI-TOF mass spectrometry (MS

  15. MicroRNA profile analysis of host cells before and after wild human rotavirus infection.

    Science.gov (United States)

    Zhou, Yan; Wu, Jinyuan; Geng, Panpan; Kui, Xiang; Xie, Yuping; Zhang, Lei; Liu, Yaling; Yin, Na; Zhang, Guangming; Yi, Shan; Li, Hongjun; Sun, Maosheng

    2016-09-01

    Rotavirus infection is an important cause of acute gastroenteritis in children, but the interaction between rotavirus and host cells is not completely understood. We isolated a wildtype (wt) rotavirus strain, ZTR-68(P [8] G1), which is derived from an infant with diarrhea in southwest China in 2010. In this study, we investigated host cellular miRNA expression profiles changes in response to ZTR-68 in early stage of infection to investigate the role of miRNAs upon rotavirus infection. Differentially expressed miRNAs were identified by deep sequencing and qRT-PCR and the function of their targets predicted by Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation. A total of 36 candidate miRNAs were identified. Comparative analysis indicated that 29 miRNAs were significantly down-regulated and 7 were up-regulated after infection. The data were provided contrasting the types of microRNAs in two different permissive cell lines (HT29 and MA104). The target assays results showed that mml-miR-7 and mml-miR-125a are involved in anti-rotavirus and virus-host interaction in host cells. These results offer clues for identifying potential candidates in vector-based antiviral strategies. J. Med. Virol. 88:1497-1510, 2016. © 2016 Wiley Periodicals, Inc. PMID:26890217

  16. A bacterial type III effector family uses the papain-like hydrolytic activity to arrest the host cell cycle

    OpenAIRE

    Yao, Qing; Cui, Jixin; Zhu, Yongqun; Wang, Guolun; Hu, Liyan; Long, Chengzu; Cao, Ran; Liu, Xinqi; Huang, Niu; Chen, She; Liu, LiPing; Shao, Feng

    2009-01-01

    Pathogenic bacteria deliver effector proteins into host cells through the type III secretion apparatus to modulate the host function. We identify a family of proteins, homologous to the type III effector Cif from enteropathogenic Escherichia coli, in pathogens including Yersinia, Photorhabdus, and Burkholderia that contain functional type III secretion systems. Like Cif, this family of proteins is capable of arresting the host cell cycle at G2/M. Structure of one of the family members, Cif ho...

  17. Host and viral factors contributing to CD8+ T cell failure in hepatitis C virus infection

    Institute of Scientific and Technical Information of China (English)

    Christoph Neumann-Haefelin; Hans Christian Spangenberg; Hubert E Blum; Robert Thimme

    2007-01-01

    Virus-specific CD8+ T cells are thought to be the major anti-viral effector cells in hepatitis C virus (HCV)infection. Indeed, viral clearance is associated with vigorous CD8+ T cell responses targeting multiple epitopes. In the chronic phase of infection, HCV-specific CD8+ T cell responses are usually weak, narrowly focused and display often functional defects regarding cytotoxicity, cytokine production, and proliferative capacity. In the last few years, different mechanisms which might contribute to the failure of HCV-specific CD8+ T cells in chronic infection have been identified,including insufficient CD4+ help, deficient CD8+ T cell differentiation, viral escape mutations, suppression by viral factors, inhibitory cytokines, inhibitory ligands, and regulatory T cells. In addition, host genetic factors such as the host's human leukocyte antigen (HLA) background may play an important role in the efficiency of the HCVspecific CD8+ T cell response and thus outcome of infection. The growing understanding of the mechanisms contributing to T cell failure and persistence of HCV infection will contribute to the development of successful immunotherapeutical and -prophylactical strategies.

  18. Alterations in cell surface area and deformability of individual human red blood cells in stored blood

    CERN Document Server

    Park, HyunJoo; Lee, SangYun; Kim, Kyoohyun; Sohn, Yong-Hak; Jang, Seongsoo; Park, YongKeun

    2015-01-01

    The functionality and viability of stored human red blood cells (RBCs) is an important clinical issue in transfusion. To systematically investigate changes in stored whole blood, the hematological properties of individual RBCs were quantified in blood samples stored for various periods with and without a preservation solution called CPDA-1. With 3-D quantitative phase imaging techniques, the optical measurements of the 3-D refractive index (RI) distributions and membrane fluctuations were done at the individual cell level. From the optical measurements, the morphological (volume, surface area and sphericity), biochemical (hemoglobin content and concentration), and mechanical parameters (dynamic membrane fluctuation) were simultaneously quantified to investigate the functionalities and their progressive alterations in stored RBCs. Our results show that the stored RBCs without CPDA-1 had a dramatic morphological transformation from discocytes to spherocytes within 2 weeks which was accompanied with significant ...

  19. Ultrastructural characteristics of nurse cell-larva complex of four species of Trichinella in several hosts

    Directory of Open Access Journals (Sweden)

    Sacchi L.

    2001-06-01

    Full Text Available The nurse cell-larva complex of nematodes of the genus Trichinella plays an Important role in the survival of the larva in decaying muscles, frequently favouring the transmission of the parasite in extreme environmental conditions. The ultrastructure of the nurse cell-larva complex in muscles from different hosts infected with T. nativa (a walrus and a polar bear, T. spiralis (horses and humans, T. pseudospiralis (a laboratory mouse and T. papuae (a laboratory mouse were examined. Analysis with transmission electron microscope showed that the typical nurse cell structure was present in all examined samples, irrespective of the species of larva, of the presence of a collagen capsule, of the age of infection and of the host species, suggesting that there exists a molecular mechanism that in the first stage of larva invasion is similar for encapsulated and non-encapsulated species.

  20. Altered Membrane Potential and Electrolyte in Sickle Cell Anemia

    Directory of Open Access Journals (Sweden)

    JK Nnodim

    2014-01-01

    Full Text Available Aim: This study has been to evaluate the level of membrane potential and electrolyte in sickle cell disease patients. Material and methods: 100 sickle cell patients in steady state ages 5 to 30 years attending General Hospital Owerri were used in the study while 100 normal subjects (HbAA were used as control. Also 30 HbSS in crisis have been involved. Results: The results obtained showed that the level of membrane potential was significantly lower in sickle cell anemia as compared to the controls. Also, the level of the electrolyte was found significantly decreased in HbSS when compared with HbAA at P<0.05. Conclusion: The membrane potential translates to energy which means that there is less energy in sickle cell disease which is linked to electrolyte imbalance. Hence people with sickle disease should be monitored closely for their electrolytes to avoid crisis.

  1. Alteration of mammalian cell metabolism by dynamic nutrient feeding

    OpenAIRE

    Zhou, Weichang; Rehm, Jutta; Europa, Anna; Hu, Wei-Shou

    1997-01-01

    The metabolism of hybridoma cells was controlled to reduce metabolic formation in fed-batch cultures by dynamically feeding a salt-free nutrient concentrate. For this purpose, on-line oxygen uptake rate (OUR) measurement was used to estimate the metabolic demand of hybridoma cells and to determine the feeding rate of a concentrated solution of salt-free DMEM/F12 medium supplemented with other medium components. The ratios among glucose, glutamine and other medium components in the feeding nut...

  2. Alterations in auxin homeostasis suppress defects in cell wall function.

    Directory of Open Access Journals (Sweden)

    Blaire J Steinwand

    Full Text Available The plant cell wall is a highly dynamic structure that changes in response to both environmental and developmental cues. It plays important roles throughout plant growth and development in determining the orientation and extent of cell expansion, providing structural support and acting as a barrier to pathogens. Despite the importance of the cell wall, the signaling pathways regulating its function are not well understood. Two partially redundant leucine-rich-repeat receptor-like kinases (LRR-RLKs, FEI1 and FEI2, regulate cell wall function in Arabidopsis thaliana roots; disruption of the FEIs results in short, swollen roots as a result of decreased cellulose synthesis. We screened for suppressors of this swollen root phenotype and identified two mutations in the putative mitochondrial pyruvate dehydrogenase E1α homolog, IAA-Alanine Resistant 4 (IAR4. Mutations in IAR4 were shown previously to disrupt auxin homeostasis and lead to reduced auxin function. We show that mutations in IAR4 suppress a subset of the fei1 fei2 phenotypes. Consistent with the hypothesis that the suppression of fei1 fei2 by iar4 is the result of reduced auxin function, disruption of the WEI8 and TAR2 genes, which decreases auxin biosynthesis, also suppresses fei1 fei2. In addition, iar4 suppresses the root swelling and accumulation of ectopic lignin phenotypes of other cell wall mutants, including procuste and cobra. Further, iar4 mutants display decreased sensitivity to the cellulose biosynthesis inhibitor isoxaben. These results establish a role for IAR4 in the regulation of cell wall function and provide evidence of crosstalk between the cell wall and auxin during cell expansion in the root.

  3. Genetic Alterations in Gliosarcoma and Giant Cell Glioblastoma.

    Science.gov (United States)

    Oh, Ji Eun; Ohta, Takashi; Nonoguchi, Naosuke; Satomi, Kaishi; Capper, David; Pierscianek, Daniela; Sure, Ulrich; Vital, Anne; Paulus, Werner; Mittelbronn, Michel; Antonelli, Manila; Kleihues, Paul; Giangaspero, Felice; Ohgaki, Hiroko

    2016-07-01

    The majority of glioblastomas develop rapidly with a short clinical history (primary glioblastoma IDH wild-type), whereas secondary glioblastomas progress from diffuse astrocytoma or anaplastic astrocytoma. IDH mutations are the genetic hallmark of secondary glioblastomas. Gliosarcomas and giant cell glioblastomas are rare histological glioblastoma variants, which usually develop rapidly. We determined the genetic patterns of 36 gliosarcomas and 19 giant cell glioblastomas. IDH1 and IDH2 mutations were absent in all 36 gliosarcomas and in 18 of 19 giant cell glioblastomas analyzed, indicating that they are histological variants of primary glioblastoma. Furthermore, LOH 10q (88%) and TERT promoter mutations (83%) were frequent in gliosarcomas. Copy number profiling using the 450k methylome array in 5 gliosarcomas revealed CDKN2A homozygous deletion (3 cases), trisomy chromosome 7 (2 cases), and monosomy chromosome 10 (2 cases). Giant cell glioblastomas had LOH 10q in 50% and LOH 19q in 42% of cases. ATRX loss was detected immunohistochemically in 19% of giant cell glioblastomas, but absent in 17 gliosarcomas. These and previous results suggest that gliosarcomas are a variant of, and genetically similar to, primary glioblastomas, except for a lack of EGFR amplification, while giant cell glioblastoma occupies a hybrid position between primary and secondary glioblastomas. PMID:26443480

  4. Donor Requirements for Regulatory T Cell Suppression of Murine Graft Versus Host Disease

    OpenAIRE

    Pierini, Antonio; Colonna, Lucrezia; Alvarez, Maite; Schneidawind, Dominik; Nishikii, Hidekazu; Baker, Jeanette; Pan, Yuqiong; Florek, Mareike; Kim, Byung-Su; Negrin, Robert S.

    2015-01-01

    Adoptive transfer of freshly isolated natural occurring CD4+CD25+FoxP3+ regulatory T cells (Treg) prevents graft versus host disease (GvHD) in several animal models and following hematopoietic cell transplantation (HCT) in clinical trials. Donor derived Treg have been mainly used as they share the same MHC with conventional CD4+ and CD8+ T cells (Tcon) that are primarily responsible for GvHD. Third-party derived Treg are a promising alternative for cellular therapy as they can be prepared in ...

  5. Receptor ganglioside content of three hosts for Sendai virus. MDBK, HeLa, and MDCK cells.

    Science.gov (United States)

    Markwell, M A; Fredman, P; Svennerholm, L

    1984-08-01

    Specific gangliosides GD1a, GT1b and GQ1b isolated from brain have been shown to function as receptors for Sendai virus by conferring susceptibility to infection when they are incorporated into receptor-deficient cells (Markwell, M.A.K., Svennerholm, L. and Paulson, J.C. (1981) Proc. Natl. Acad. Sci. USA 78, 5406-5410). The endogenous gangliosides of three commonly used hosts for Sendai virus: MDBK, HeLa, and MDCK cells were analyzed to determine the amount and type of receptor gangliosides present. In all three cell lines, GM3 was the major ganglioside component. The presence of GM1, GD1a and the more complex homologs of the gangliotetraose series was also established. In cell lines derived from normal tissue, MDBK and MDCK cells, gangliosides contributed 47-65% of the total sialic acid. In HeLa cells, gangliosides contributed substantially less (17% of the total sialic acid). The ganglioside content of each cell line was shown not to be immutable but instead to depend on the state of differentiation, passage number, and surface the cells were grown on. Thus, the ganglioside concentration of undifferentiated MDCK cells was found to be substantially greater than that of MDBK or HeLa cells, but decreased as the MDCK cells underwent differentiation. Changes in culture conditions that were shown to decrease the receptor ganglioside content of the cells resulted in a corresponding decrease in susceptibility to infection. The endogenous oligosialogangliosides present in susceptible host cells were shown to function as receptors for Sendai virus.

  6. Lipid body accumulation alters calcium signaling dynamics in immune cells.

    Science.gov (United States)

    Greineisen, William E; Speck, Mark; Shimoda, Lori M N; Sung, Carl; Phan, Nolwenn; Maaetoft-Udsen, Kristina; Stokes, Alexander J; Turner, Helen

    2014-09-01

    There is well-established variability in the numbers of lipid bodies (LB) in macrophages, eosinophils, and neutrophils. Similarly to the steatosis observed in adipocytes and hepatocytes during hyperinsulinemia and nutrient overload, immune cell LB hyper-accumulate in response to bacterial and parasitic infection and inflammatory presentations. Recently we described that hyperinsulinemia, both in vitro and in vivo, drives steatosis and phenotypic changes in primary and transformed mast cells and basophils. LB reach high numbers in these steatotic cytosols, and here we propose that they could dramatically impact the transcytoplasmic signaling pathways. We compared calcium release and influx responses at the population and single cell level in normal and steatotic model mast cells. At the population level, all aspects of FcɛRI-dependent calcium mobilization, as well as activation of calcium-dependent downstream signaling targets such as NFATC1 phosphorylation are suppressed. At the single cell level, we demonstrate that LB are both sources and sinks of calcium following FcɛRI cross-linking. Unbiased analysis of the impact of the presence of LB on the rate of trans-cytoplasmic calcium signals suggest that LB enrichment accelerates calcium propagation, which may reflect a Bernoulli effect. LB abundance thus impacts this fundamental signaling pathway and its downstream targets.

  7. Selection of mutant Chinese hamster ovary cells altered glycoproteins by means of tritiated fucose suicide.

    OpenAIRE

    Hirschberg, C B; Baker, R.M.; Perez, M.; Spencer, L A; Watson, D

    1981-01-01

    Mutant Chinese hamster ovary cells altered in glycoproteins have been isolated by selecting for ability to survive exposure to [6-3H]fucose. Mutagenized wild-type cells were permitted to incorporate [3H]fucose to approximately 1 cpm of trichloroacetic acid-insoluble radioactivity per cell and then frozen for several days to accumulate radiation damage. The overall viability of the population was reduced by 5- to 50-fold. Four consecutive selection cycles were carried out. The surviving cells ...

  8. Bactericidal Antibiotics Increase Hydroxyphenyl Fluorescein Signal by Altering Cell Morphology

    DEFF Research Database (Denmark)

    Paulander, Wilhelm; Wang, Ying; Folkesson, Sven Anders;

    2014-01-01

    It was recently proposed that for bactericidal antibiotics a common killing mechanism contributes to lethality involving indirect stimulation of hydroxyl radical (OH center dot) formation. Flow cytometric detection of OH center dot by hydroxyphenyl fluorescein (HPF) probe oxidation was used...... to support this hypothesis. Here we show that increased HPF signals in antibiotics-exposed bacterial cells are explained by fluorescence associated with increased cell size, and do not reflect reactive oxygen species (ROS) concentration. Independently of antibiotics, increased fluorescence was seen...... for elongated cells expressing the oxidative insensitive green fluorescent protein (GFP). Although our data question the role of ROS in lethality of antibiotics other research approaches point to important interplays between basic bacterial metabolism and antibiotic susceptibility. To underpin...

  9. Alphavirus Infection: Host Cell Shut-Off and Inhibition of Antiviral Responses

    Directory of Open Access Journals (Sweden)

    Jelke J. Fros

    2016-06-01

    Full Text Available Alphaviruses cause debilitating disease in humans and animals and are transmitted by blood-feeding arthropods, typically mosquitoes. With a traditional focus on two models, Sindbis virus and Semliki Forest virus, alphavirus research has significantly intensified in the last decade partly due to the re-emergence and dramatic expansion of chikungunya virus in Asia, Europe, and the Americas. As a consequence, alphavirus–host interactions are now understood in much more molecular detail, and important novel mechanisms have been elucidated. It has become clear that alphaviruses not only cause a general host shut-off in infected vertebrate cells, but also specifically suppress different host antiviral pathways using their viral nonstructural proteins, nsP2 and nsP3. Here we review the current state of the art of alphavirus host cell shut-off of viral transcription and translation, and describe recent insights in viral subversion of interferon induction and signaling, the unfolded protein response, and stress granule assembly.

  10. Immunoregulatory Effects Triggered by Lactic Acid Bacteria Exopolysaccharides: New Insights into Molecular Interactions with Host Cells

    Directory of Open Access Journals (Sweden)

    Jonathan Laiño

    2016-08-01

    Full Text Available Researchers have demonstrated that lactic acid bacteria (LAB with immunomodulatory capabilities (immunobiotics exert their beneficial effects through several molecules, including cell wall, peptidoglycan, and exopolysaccharides (EPS, that are able to interact with specific host cell receptors. EPS from LAB show a wide heterogeneity in its composition, meaning that biological properties depend on the strain and. therefore, only a part of the mechanism of action has been elucidated for these molecules. In this review, we summarize the current knowledge of the health-promoting actions of EPS from LAB with special focus on their immunoregulatory actions. In addition, we describe our studies using porcine intestinal epithelial cells (PIE cells as a model to evaluate the molecular interactions of EPS from two immunobiotic LAB strains and the host cells. Our studies showed that EPS from immunobiotic LAB have anti-inflammatory capacities in PIE cells since they are able to reduce the production of inflammatory cytokines in cells challenged with the Toll-like receptor (TLR-4-agonist lipopolysaccharide. The effects of EPS were dependent on TLR2, TLR4, and negative regulators of TLR signaling. We also reported that the radioprotective 105 (RP105/MD1 complex, a member of the TLR family, is partially involved in the immunoregulatory effects of the EPS from LAB. Our work described, for the first time, that LAB and their EPS reduce inflammation in intestinal epithelial cells in a RP105/MD1-dependent manner. A continuing challenge for the future is to reveal more effector-receptor relationships in immunobiotic-host interactions that contribute to the beneficial effects of these bacteria on mucosal immune homeostasis. A detailed molecular understanding should lead to a more rational use of immunobiotics in general, and their EPS in particular, as efficient prevention and therapies for specific immune-related disorders in humans and animals.

  11. Lipid rafts, caveolae, caveolin-1, and entry by Chlamydiae into host cells.

    Science.gov (United States)

    Stuart, Elizabeth S; Webley, Wilmore C; Norkin, Leonard C

    2003-07-01

    Obligate intracellular bacterial pathogens of the genus Chlamydia are reported to enter host cells by both clathrin-dependent and clathrin-independent processes. C. trachomatis serovar K recently was shown to enter cells via caveolae-like lipid raft domains. We asked here how widespread raft-mediated entry might be among the Chlamydia. We show that C. pneumoniae, an important cause of respiratory infections in humans that additionally is associated with cardiovascular disease, and C. psittaci, an important pathogen in domestic mammals and birds that also infects humans, each enter host cells via cholesterol-rich lipid raft microdomains. Further, we show that C. trachomatis serovars E and F also use these domains to enter host cells. The involvement of these membrane domains in the entry of these organisms was indicated by the sensitivity of their entry to the raft-disrupting agents Nystatin and filipin, and by their intracellular association with caveolin-1, a 22-kDa protein associated with the formation of caveolae in rafts. In contrast, caveolin-marked lipid raft domains do not mediate entry of C. trachomatis serovars A, 36B, and C, nor of LGV serovar L2 and MoPn. Finally, we show that entry of each of these chlamydial strains is independent of cellular expression of caveolin-1. Thus, entry via the Nystatin and filipin-sensitive pathway is dependent on lipid rafts containing cholesterol, rather than invaginated caveolae per se.

  12. Nanomimics of host cell membranes block invasion and expose invasive malaria parasites.

    Science.gov (United States)

    Najer, Adrian; Wu, Dalin; Bieri, Andrej; Brand, Françoise; Palivan, Cornelia G; Beck, Hans-Peter; Meier, Wolfgang

    2014-12-23

    The fight against most infectious diseases, including malaria, is often hampered by the emergence of drug resistance and lack or limited efficacies of vaccines. Therefore, new drugs, vaccines, or other strategies to control these diseases are needed. Here, we present an innovative nanotechnological strategy in which the nanostructure itself represents the active substance with no necessity to release compounds to attain therapeutic effect and which might act in a drug- and vaccine-like dual function. Invasion of Plasmodium falciparum parasites into red blood cells was selected as a biological model for the initial validation of this approach. Stable nanomimics-polymersomes presenting receptors required for parasite attachment to host cells-were designed to efficiently interrupt the life cycle of the parasite by inhibiting invasion. A simple way to build nanomimics without postformation modifications was established. First, a block copolymer of the receptor with a hydrophobic polymer was synthesized and then mixed with a polymersome-forming block copolymer. The resulting nanomimics bound parasite-derived ligands involved in the initial attachment to host cells and they efficiently blocked reinvasion of malaria parasites after their egress from host cells in vitro. They exhibited efficacies of more than 2 orders of magnitude higher than the soluble form of the receptor, which can be explained by multivalent interactions of several receptors on one nanomimic with multiple ligands on the infective parasite. In the future, our strategy might offer interesting treatment options for severe malaria or a way to modulate the immune response. PMID:25435059

  13. Internalization of components of the host cell plasma membrane during infection by Trypanosoma cruzi

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    Carvalho TMU

    1999-01-01

    Full Text Available Epimastigote and trypomastigote forms of Trypanosoma cruzi attach to the macrophage surface and are internalized with the formation of a membrane bounded vacuole, known as the parasitophorous vacuole (PV. In order to determine if components of the host cell membrane are internalized during formation of the PV we labeled the macrophage surface with fluorescent probes for proteins, lipids and sialic acid residues and then allowed the labeled cells to interact with the parasites. The interaction process was interrupted after 1 hr at 37ºC and the distribution of the probes analyzed by confocal laser scanning microscopy. During attachment of the parasites to the macrophage surface an intense labeling of the attachment regions was observed. Subsequently labeling of the membrane lining the parasitophorous vacuole containing epimastigote and trypomastigote forms was seen. Labeling was not uniform, with regions of intense and light or no labeling. The results obtained show that host cell membrane lipids, proteins and sialoglycoconjugates contribute to the formation of the membrane lining the PV containing epimastigote and trypomastigote T. cruzi forms. Lysosomes of the host cell may participate in the process of PV membrane formation.

  14. Concerted action of two formins in gliding motility and host cell invasion by Toxoplasma gondii.

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    Wassim Daher

    Full Text Available The invasive forms of apicomplexan parasites share a conserved form of gliding motility that powers parasite migration across biological barriers, host cell invasion and egress from infected cells. Previous studies have established that the duration and direction of gliding motility are determined by actin polymerization; however, regulators of actin dynamics in apicomplexans remain poorly characterized. In the absence of a complete ARP2/3 complex, the formin homology 2 domain containing proteins and the accessory protein profilin are presumed to orchestrate actin polymerization during host cell invasion. Here, we have undertaken the biochemical and functional characterization of two Toxoplasma gondii formins and established that they act in concert as actin nucleators during invasion. The importance of TgFRM1 for parasite motility has been assessed by conditional gene disruption. The contribution of each formin individually and jointly was revealed by an approach based upon the expression of dominant mutants with modified FH2 domains impaired in actin binding but still able to dimerize with their respective endogenous formin. These mutated FH2 domains were fused to the ligand-controlled destabilization domain (DD-FKBP to achieve conditional expression. This strategy proved unique in identifying the non-redundant and critical roles of both formins in invasion. These findings provide new insights into how controlled actin polymerization drives the directional movement required for productive penetration of parasites into host cells.

  15. Characterisation of cell death inducing Phytophthora capsici CRN effectors suggests diverse activities in the host nucleus

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    Remco eStam

    2013-10-01

    Full Text Available Plant-Microbe interactions are complex associations that feature recognition of Pathogen Associated Molecular Patterns by the plant immune system and dampening of subsequent responses by pathogen encoded secreted effectors. With large effector repertoires now identified in a range of sequenced microbial genomes, much attention centres on understanding their roles in immunity or disease. These studies not only allow identification of pathogen virulence factors and strategies, they also provide an important molecular toolset suited for studying immunity in plants. The Phytophthora intracellular effector repertoire encodes a large class of proteins that translocate into host cells and exclusively target the host nucleus. Recent functional studies have implicated the CRN protein family as an important class of diverse effectors that target distinct subnuclear compartments and modify host cell signalling. Here, we characterised three necrosis inducing CRNs and show that there are differences in the levels of cell death. We show that only expression of CRN20_624 has an additive effect on PAMP induced cell death but not AVR3a induced ETI. Given their distinctive phenotypes, we assessed localisation of each CRN with a set of nuclear markers and found clear differences in CRN subnuclear distribution patterns. These assays also revealed that expression of CRN83_152 leads to a distinct change in nuclear chromatin organisation, suggesting a distinct series of events that leads to cell death upon over-expression. Taken together, our results suggest diverse functions carried by CRN C-termini, which can be exploited to identify novel processes that take place in the host nucleus and are required for immunity or susceptibility.

  16. Immunosuppressive CD71+ erythroid cells compromise neonatal host defence against infection

    Science.gov (United States)

    Elahi, Shokrollah; Ertelt, James M.; Kinder, Jeremy M.; Jiang, Tony T.; Zhang, Xuzhe; Xin, Lijun; Chaturvedi, Vandana; Strong, Beverly S.; Qualls, Joseph E.; Steinbrecher, Kris A.; Kalfa, Theodosia A.; Shaaban, Aimen F.; Way, Sing Sing

    2013-12-01

    Newborn infants are highly susceptible to infection. This defect in host defence has generally been ascribed to the immaturity of neonatal immune cells; however, the degree of hyporesponsiveness is highly variable and depends on the stimulation conditions. These discordant responses illustrate the need for a more unified explanation for why immunity is compromised in neonates. Here we show that physiologically enriched CD71+ erythroid cells in neonatal mice and human cord blood have distinctive immunosuppressive properties. The production of innate immune protective cytokines by adult cells is diminished after transfer to neonatal mice or after co-culture with neonatal splenocytes. Neonatal CD71+ cells express the enzyme arginase-2, and arginase activity is essential for the immunosuppressive properties of these cells because molecular inhibition of this enzyme or supplementation with L-arginine overrides immunosuppression. In addition, the ablation of CD71+ cells in neonatal mice, or the decline in number of these cells as postnatal development progresses parallels the loss of suppression, and restored resistance to the perinatal pathogens Listeria monocytogenes and Escherichia coli. However, CD71+ cell-mediated susceptibility to infection is counterbalanced by CD71+ cell-mediated protection against aberrant immune cell activation in the intestine, where colonization with commensal microorganisms occurs swiftly after parturition. Conversely, circumventing such colonization by using antimicrobials or gnotobiotic germ-free mice overrides these protective benefits. Thus, CD71+ cells quench the excessive inflammation induced by abrupt colonization with commensal microorganisms after parturition. This finding challenges the idea that the susceptibility of neonates to infection reflects immune-cell-intrinsic defects and instead highlights processes that are developmentally more essential and inadvertently mitigate innate immune protection. We anticipate that these

  17. Malaria parasite interactions with the human host

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    Pouniotis D

    2004-01-01

    Full Text Available The interaction between the malaria parasite and the human host involves a number of interactions that result in the parasite evading the human immune system. Since the stages of the malaria lifecycle are complex, this allows the use of various immune evasion strategies by the malaria parasite and has major implications in the development of a vaccine for malaria endemic areas. The present review highlights key host:parasite interactions. Plasmodia puts selection pressure on human gene frequencies, and studies into host genetic factors such as the Duffy blood group and sickle cell anaemia offer insight into the host- parasite relationship. In addition, parasite interactions with the different effector arms of the immune system can result in altered peptide ligand (APL antagonism which alters the immune response from a pro- to an anti-inflammatory T cell response. Recent insights into the interaction between professional antigen presenting cells, dendritic cells (DCs, and malaria parasites is discussed in detail.

  18. Transfected parvalbumin alters calcium homeostasis in teratocarcinoma PCC7 cells

    DEFF Research Database (Denmark)

    Müller, B K; Kabos, P; Belhage, B;

    1996-01-01

    Indirect evidence supports a protective role of some EF-hand calcium-binding proteins against calcium-induced neurotoxicity. Little is known about how these proteins influence cytosolic calcium levels. After cloning the parvalbumin cDNA into an expression vector, teratocarcinoma cells (PCC7) were...

  19. Lowering extracellular chloride concentration alters outer hair cell shape.

    Science.gov (United States)

    Cecola, R P; Bobbin, R P

    1992-08-01

    In general, increasing external K+ concentration, as well as exposure to hypotonic medium, induces a shortening of outer hair cells (OHCs) accompanied by an increase in width and volume. One possible mechanism suggested for these changes is a movement of Cl- and/or water across the cell membrane. We therefore examined the role of Cl- in OHC volume maintenance by testing the effect of decreasing extracellular Cl- concentration on OHC length and shape. In addition, the effect of hypotonic medium was examined. OHCs were isolated from guinea pig cochleae, mechanically dissociated and dispersed, and placed in a modified Hanks balanced salt solution (HBS). Exposing the cells to a Cl(-)-free HBS produced an initial shortening, which was rapidly followed by an increase in length. After about 9 min of exposure to Cl(-)-free HBS, the cells appeared to lose all water and collapsed. Upon return to normal HBS, the OHCs returned to their normal shape. We speculate that the collapse of the OHCs may be due to the loss of intracellular Cl-, which, in turn, resulted in the loss of intracellular K+ and water. The results indicate that Cl- contributes greatly to the maintenance of OHC volume. In addition, we confirmed that isolated OHCs swell in hypotonic medium and maintain their swollen state until returned to normal medium. The mechanism for maintenance of the swollen state is unknown.

  20. Effect of lactoferrin protein on red blood cells and macrophages: mechanism of parasite–host interaction

    Directory of Open Access Journals (Sweden)

    An

    2015-07-01

    Full Text Available Namrata Anand,1 Rupinder K Kanwar,2 Mohan Lal Dubey,1 R K Vahishta,3 Rakesh Sehgal,1,* Anita K Verma,4 Jagat R Kanwar2,*1Department of Medical Parasitology, Postgraduate Institute of Medical Education and Research, Chandigarh, India; 2Nanomedicine Laboratory of Immunology and Molecular Biomedical Research, School of Medicine, Molecular and Medical Research Strategic Research Centre, Faculty of Health, Deakin University, Geelong, VIC, Australia; 3Department of Histopathology, Postgraduate Institute of Medical Education and Research, Chandigarh, 4Nanobiotech Laboratory, Department of Zoology, Kirorimal College, University of Delhi, Delhi, India*These authors contributed equally to this workBackground: Lactoferrin is a natural multifunctional protein known to have antitumor, antimicrobial, and anti-inflammatory activity. Apart from its antimicrobial effects, lactoferrin is known to boost the immune response by enhancing antioxidants. Lactoferrin exists in various forms depending on its iron saturation. The present study was done to observe the effect of lactoferrin, isolated from bovine and buffalo colostrum, on red blood cells (RBCs and macrophages (human monocytic cell line-derived macrophages THP1 cells.Methods: Lactoferrin obtained from both species and in different iron saturation forms were used in the present study, and treatment of host cells were given with different forms of lactoferrin at different concentrations. These treated host cells were used for various studies, including morphometric analysis, viability by MTT assay, survivin gene expression, production of reactive oxygen species, phagocytic properties, invasion assay, and Toll-like receptor-4, Toll-like receptor-9, and MDR1 expression, to investigate the interaction between lactoferrin and host cells and the possible mechanism of action with regard to parasitic infections.Results: The mechanism of interaction between host cells and lactoferrin have shown various aspects of gene

  1. From microbe to man: the role of microbial short chain fatty acid metabolites in host cell biology.

    Science.gov (United States)

    Natarajan, Niranjana; Pluznick, Jennifer L

    2014-12-01

    Recent studies have highlighted a myriad of ways in which the activity and composition of the gut microbiota can affect the host organism. A primary way in which the gut microbiota affect host physiology is by the production of metabolites, such as short-chain fatty acids (SCFAs), which are subsequently absorbed into the bloodstream of the host. Although recent studies have begun to unravel the ways in which gut microbial SCFAs affect host physiology, less is understood regarding the underlying cell biological mechanisms. In this review, we will outline the known receptors and transporters for SCFAs, and review what is known about the cell biological effects of microbial SCFAs. PMID:25273884

  2. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture

    Science.gov (United States)

    Lestard, Nathalia R.

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  3. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture.

    Science.gov (United States)

    Lestard, Nathalia R; Capella, Marcia A M

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  4. Host epithelial cell invasion by Campylobacter jejuni: trigger or zipper mechanism?

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    Tadhg eÓ Cróinín

    2012-03-01

    Full Text Available Campylobacter jejuni, a spiral-shaped Gram-negative pathogen, is a highly frequent cause of gastrointestinal foodborne illness in humans worldwide. Clinical outcome of C. jejuni infections ranges from mild to severe diarrheal disease, and some other complications including reactive arthritis and Guillain–Barré syndrome. This review article highlights various C. jejuni pathogenicity factors, host cell determinants and proposed signaling mechanisms involved in human host cell invasion and their potential role in the development of C. jejuni-mediated disease. A model is presented which outlines the various important interactions of C. jejuni with the intestinal epithelium, and we discuss the pro’s and con’s for the zipper over the trigger mechanism of invasion. Future work should clarify the contradictory role of some previously identified factors, and should identify and characterize novel virulence determinants, which are crucial to provide fresh insights into the diversity of strategies employed by this pathogen to cause disease.

  5. Equine herpesvirus type 1 modulates inflammatory host immune response genes in equine endothelial cells.

    Science.gov (United States)

    Johnstone, Stephanie; Barsova, Jekaterina; Campos, Isabel; Frampton, Arthur R

    2016-08-30

    Equine herpesvirus myeloencephalopathy (EHM), a disease caused by equine herpesvirus type 1 (EHV-1), is characterized by severe inflammation, thrombosis, and hypoxia in central nervous system (CNS) endothelial cells, which can result in a spectrum of clinical signs including urinary incontinence, ataxia, and paralysis. Strains of EHV-1 that contain a single point mutation within the viral DNA polymerase (nucleotide A2254>G2254: amino acid N752→D752) are isolated from EHM afflicted horses at higher frequencies than EHV-1 strains that do not harbor this mutation. Due to the correlation between the DNA Pol mutation and EHM disease, EHV-1 strains that contain the mutation have been designated as neurologic. In this study, we measured virus replication, cell to cell spread efficacy, and host inflammatory responses in equine endothelial cells infected with 12 different strains of EHV-1. Two strains, T953 (Ohio 2003) (neurologic) and Kentucky A (KyA) (non-neurologic), have well described disease phenotypes while the remaining strains used in this study are classified as neurologic or non-neurologic based solely on the presence or absence of the DNA pol mutation, respectively. Results show that the neurologic strains do not replicate better or spread more efficiently in endothelial cells. Also, the majority of the host inflammatory genes were modulated similarly regardless of EHV-1 genotype. Analyses of host gene expression showed that a subset of pro-inflammatory cytokines, including the CXCR3 ligands CXCL9, CXCL10, and CXCL11, as well as CCL5, IL-6 and TNF-α were consistently up-regulated in endothelial cells infected with each EHV-1 strain. The identification of specific pro-inflammatory cytokines in endothelial cells that are modulated by EHV-1 provides further insight into the factors that contribute to the immunopathology observed after infection and may also reveal new targets for disease intervention. PMID:27527764

  6. Is the optimal pH for membrane fusion in host cells by avian influenza viruses related to host range and pathogenicity?

    Science.gov (United States)

    Okamatsu, Masatoshi; Motohashi, Yurie; Hiono, Takahiro; Tamura, Tomokazu; Nagaya, Kazuki; Matsuno, Keita; Sakoda, Yoshihiro; Kida, Hiroshi

    2016-08-01

    Influenza viruses isolated from wild ducks do not replicate in chickens. This fact is not explained solely by the receptor specificity of the hemagglutinin (HA) from such viruses for target host cells. To investigate this restriction in host range, the fusion activities of HA molecules from duck and chicken influenza viruses were examined. Influenza viruses A/duck/Mongolia/54/2001 (H5N2) (Dk/MNG) and A/chicken/Ibaraki/1/2005 (H5N2) (Ck/IBR), which replicate only in their primary hosts, were used. The optimal pH for membrane fusion of Ck/IBR was 5.9, higher than that of Dk/MNG at 4.9. To assess the relationship between the optimal pH for fusion and the host range of avian influenza viruses, the optimal pH for fusion of 55 influenza virus strains isolated from ducks and chickens was examined. No correlation was found between the host range and optimal pH for membrane fusion by the viruses, and this finding applied also to the H5N1 highly pathogenic avian influenza viruses. The optimal pH for membrane fusion for avian influenza viruses was shown to not necessarily be correlated with their host range or pathogenicity in ducks and chickens. PMID:27231009

  7. NK and NKT Cell Depletion Alters the Outcome of Experimental Pneumococcal Pneumonia: Relationship with Regulation of Interferon-γ Production

    Directory of Open Access Journals (Sweden)

    Eirini Christaki

    2015-01-01

    Full Text Available Background. Natural killer (NK and natural killer T (NKT cells contribute to the innate host defense but their role in bacterial sepsis remains controversial. Methods. C57BL/6 mice were infected intratracheally with 5 × 105 cfu of Streptococcus pneumoniae. Animals were divided into sham group (Sham; pretreated with isotype control antibody (CON group; pretreated with anti-asialo GM1 antibody (NKd group; and pretreated with anti-CD1d monoclonal antibody (NKTd group before bacterial challenge. Serum and tissue samples were analyzed for bacterial load, cytokine levels, splenocyte apoptosis rates, and cell characteristics by flow cytometry. Splenocyte miRNA expression was also analyzed and survival was assessed. Results. NK cell depletion prolonged survival. Upon inhibition of NKT cell activation, spleen NK (CD3−/NK1.1+ cells increased compared to all other groups. Inhibition of NKT cell activation led to higher bacterial loads and increased levels of serum and splenocyte IFN-γ. Splenocyte miRNA analysis showed that miR-200c and miR-29a were downregulated, while miR-125a-5p was upregulated, in anti-CD1d treated animals. These changes were moderate after NK cell depletion. Conclusions. NK cells appear to contribute to mortality in pneumococcal pneumonia. Inhibition of NKT cell activation resulted in an increase in spleen NK (CD3−/NK1.1+ cells and a higher IFN-γ production, while altering splenocyte miRNA expression.

  8. Butachlor, a suspected carcinogen, alters growth and transformation characteristics of mouse liver cells.

    Science.gov (United States)

    Ou, Y H; Chung, P C; Chang, Y C; Ngo, F Q; Hsu, K Y; Chen, F D

    2000-12-01

    Butachlor is a widely used herbicide in Asia and South America. Previous investigations have indicated that it is a suspected carcinogen. To understand more about the biological effects of butachlor on cultured cells and the mechanism(s) of its carcinogenicity, we studied the alteration of the growth characteristics that was induced by butachlor in normal mouse liver cells (BNL CL2). This study demonstrates that butachlor decreases the population-doubling time of BNL CL2 cells, suggesting that it stimulates cell proliferation. To support this finding, a thymidine incorporation assay was conducted and a similar result that butachlor stimulates cell proliferation was elucidated. In addition, we show that butachlor increases the saturation density of the BNL CL2 cells. When combined with the tumor initiator N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), butachlor transforms cells efficiently, as demonstrated by loss of contact inhibition. These findings indicate that butachlor alters the growth characteristics of BNL CL2 cells and suggest that butachlor may induce malignant transformation through stimulation of cell proliferation, alteration of cell cycle regulation, and suppression of cell density-dependent inhibition of proliferation.

  9. Simvastatin inhibits Staphylococcus aureus host cell invasion through modulation of isoprenoid intermediates

    OpenAIRE

    Horn, Mary P.; Knecht, Sharmon M.; Rushing, Frances L.; Birdsong, Julie; Siddall, C. Parker; Johnson, Charron M.; Abraham, Terri N.; Brown, Amy; Volk, Catherine B.; Gammon, Kelly; Bishop, Derron L.; McKillip, John L.; McDowell, Susan A.

    2008-01-01

    Patients on a statin regimen are at a decreased risk of death due to bacterial sepsis. We have found that protection by simvastatin includes the inhibition of host cell invasion by Staphylococcus aureus, the most common etiologic agent of sepsis. Inhibition was due in part to depletion of isoprenoid intermediates within the cholesterol biosynthesis pathway and led to the cytosolic accumulation of the small-guanosine triphosphatases (GTPases) CDC42, Rac, and RhoB. Actin stress fiber disassembl...

  10. Influenza A virus survival in water is influenced by the origin species of the host cell

    OpenAIRE

    Shigematsu, Sayuri; Dublineau, Amélie; Sawoo, Olivier; Batéjat, Christophe; Matsuyama, Toshifumi; Leclercq, India; Manuguerra, Jean-Claude

    2014-01-01

    Background Influenza A viruses have an envelope made of a lipid bilayer and two surface glycoproteins, the hemagglutinin and the neuraminidase. The structure of the virus is directly dependent on the genetic makeup of the viral genome except the glycosylation moieties and the composition of the lipid bilayer. They both depend on the host cell and are in direct contact with the environment, such as air or water. Virus survival is important for virus transmission from contaminated waters in the...

  11. Influenza A virus survival in water is influenced by the origin species of the host cell

    OpenAIRE

    Shigematsu, Sayuri; Dublineau, Amélie; Sawoo, Olivier; Batéjat, Christophe; Matsuyama, Toshifumi; Leclercq, India; Manuguerra, Jean-Claude

    2014-01-01

    Background: Influenza A viruses have an envelope made of a lipid bilayer and two surface glycoproteins, the hemagglutinin and the neuraminidase. The structure of the virus is directly dependent on the genetic makeup of the viral genome except the glycosylation moieties and the composition of the lipid bilayer. They both depend on the host cell and are in direct contact with the environment, such as air or water. Virus survival is important for virus transmission from contaminated waters in th...

  12. Inhibition of gold nanoparticles (AuNPs) on pathogenic biofilm formation and invasion to host cells

    OpenAIRE

    Qilin Yu; Jianrong Li; Yueqi Zhang; Yufan Wang; Lu Liu; Mingchun Li

    2016-01-01

    Owing to the growing infectious diseases caused by eukaryotic and prokaryotic pathogens, it is urgent to develop novel antimicrobial agents against clinical pathogenic infections. Biofilm formation and invasion into the host cells are vital processes during pathogenic colonization and infection. In this study, we tested the inhibitory effect of Au nanoparticles (AuNPs) on pathogenic growth, biofilm formation and invasion. Interestingly, although the synthesized AuNPs had no significant toxici...

  13. Use of Genetically Altered Stem Cells for the Treatment of Huntington’s Disease

    Directory of Open Access Journals (Sweden)

    Andrew T. Crane

    2014-03-01

    Full Text Available Transplantation of stem cells for the treatment of Huntington’s disease (HD garnered much attention prior to the turn of the century. Several studies using mesenchymal stem cells (MSCs have indicated that these cells have enormous therapeutic potential in HD and other disorders. Advantages of using MSCs for cell therapies include their ease of isolation, rapid propagation in culture, and favorable immunomodulatory profiles. However, the lack of consistent neuronal differentiation of transplanted MSCs has limited their therapeutic efficacy to slowing the progression of HD-like symptoms in animal models of HD. The use of MSCs which have been genetically altered to overexpress brain derived neurotrophic factor to enhance support of surviving cells in a rodent model of HD provides proof-of-principle that these cells may provide such prophylactic benefits. New techniques that may prove useful for cell replacement therapies in HD include the use of genetically altering fate-restricted cells to produce induced pluripotent stem cells (iPSCs. These iPSCs appear to have certain advantages over the use of embryonic stem cells, including being readily available, easy to obtain, less evidence of tumor formation, and a reduced immune response following their transplantation. Recently, transplants of iPSCs have shown to differentiate into region-specific neurons in an animal model of HD. The overall successes of using genetically altered stem cells for reducing neuropathological and behavioral deficits in rodent models of HD suggest that these approaches have considerable potential for clinical use. However, the choice of what type of genetically altered stem cell to use for transplantation is dependent on the stage of HD and whether the end-goal is preserving endogenous neurons in early-stage HD, or replacing the lost neurons in late-stage HD. This review will discuss the current state of stem cell technology for treating the different stages of HD and

  14. Natural variation in the promoter of the gene encoding the Mga regulator alters host-pathogen interactions in group a Streptococcus carrier strains.

    Science.gov (United States)

    Flores, Anthony R; Olsen, Randall J; Wunsche, Andrea; Kumaraswami, Muthiah; Shelburne, Samuel A; Carroll, Ronan K; Musser, James M

    2013-11-01

    Humans commonly carry pathogenic bacteria asymptomatically, but the molecular factors underlying microbial asymptomatic carriage are poorly understood. We previously reported that two epidemiologically unassociated serotype M3 group A Streptococcus (GAS) carrier strains had an identical 12-bp deletion in the promoter of the gene encoding Mga, a global positive gene regulator. Herein, we report on studies designed to test the hypothesis that the identified 12-bp deletion in the mga promoter alters GAS virulence, thereby potentially contributing to the asymptomatic carrier phenotype. Using allelic exchange, we introduced the variant promoter into a serotype M3 invasive strain and the wild-type promoter into an asymptomatic carrier strain. Compared to strains with the wild-type mga promoter, we discovered that strains containing the promoter with the 12-bp deletion produced significantly fewer mga and Mga-regulated gene transcripts. Consistent with decreased mga transcripts, strains containing the variant mga promoter were also significantly less virulent in in vivo and ex vivo models of GAS disease. Further, we provide evidence that the pleiotropic regulator protein CodY binds to the mga promoter and that the 12-bp deletion in the mga promoter reduces CodY-mediated mga transcription. We conclude that the naturally occurring 12-bp deletion in the mga promoter significantly alters the pathogen-host interaction of these asymptomatic carrier strains. Our findings provide new insight into the molecular basis of the carrier state of an important human pathogen. PMID:23980109

  15. Role of Protein Glycosylation in Candida parapsilosis Cell Wall Integrity and Host Interaction

    Science.gov (United States)

    Pérez-García, Luis A.; Csonka, Katalin; Flores-Carreón, Arturo; Estrada-Mata, Eine; Mellado-Mojica, Erika; Németh, Tibor; López-Ramírez, Luz A.; Toth, Renata; López, Mercedes G.; Vizler, Csaba; Marton, Annamaria; Tóth, Adél; Nosanchuk, Joshua D.; Gácser, Attila; Mora-Montes, Héctor M.

    2016-01-01

    Candida parapsilosis is an important, emerging opportunistic fungal pathogen. Highly mannosylated fungal cell wall proteins are initial contact points with host immune systems. In Candida albicans, Och1 is a Golgi α1,6-mannosyltransferase that plays a key role in the elaboration of the N-linked mannan outer chain. Here, we disrupted C. parapsilosis OCH1 to gain insights into the contribution of N-linked mannosylation to cell fitness and to interactions with immune cells. Loss of Och1 in C. parapsilosis resulted in cellular aggregation, failure of morphogenesis, enhanced susceptibility to cell wall perturbing agents and defects in wall composition. We removed the cell wall O-linked mannans by β-elimination, and assessed the relevance of mannans during interaction with human monocytes. Results indicated that O-linked mannans are important for IL-1β stimulation in a dectin-1 and TLR4-dependent pathway; whereas both, N- and O-linked mannans are equally important ligands for TNFα and IL-6 stimulation, but neither is involved in IL-10 production. Furthermore, mice infected with C. parapsilosis och1Δ null mutant cells had significantly lower fungal burdens compared to wild-type (WT)-challenged counterparts. Therefore, our data are the first to demonstrate that C. parapsilosis N- and O-linked mannans have different roles in host interactions than those reported for C. albicans. PMID:27014229

  16. The bacterium endosymbiont of Crithidia deanei undergoes coordinated division with the host cell nucleus.

    Directory of Open Access Journals (Sweden)

    Maria Cristina Machado Motta

    Full Text Available In trypanosomatids, cell division involves morphological changes and requires coordinated replication and segregation of the nucleus, kinetoplast and flagellum. In endosymbiont-containing trypanosomatids, like Crithidia deanei, this process is more complex, as each daughter cell contains only a single symbiotic bacterium, indicating that the prokaryote must replicate synchronically with the host protozoan. In this study, we used light and electron microscopy combined with three-dimensional reconstruction approaches to observe the endosymbiont shape and division during C. deanei cell cycle. We found that the bacterium replicates before the basal body and kinetoplast segregations and that the nucleus is the last organelle to divide, before cytokinesis. In addition, the endosymbiont is usually found close to the host cell nucleus, presenting different shapes during the protozoan cell cycle. Considering that the endosymbiosis in trypanosomatids is a mutualistic relationship, which resembles organelle acquisition during evolution, these findings establish an excellent model for the understanding of mechanisms related with the establishment of organelles in eukaryotic cells.

  17. T-Cell Costimulatory Molecules in Acute-Graft-Versus Host Disease: Therapeutic Implications

    Directory of Open Access Journals (Sweden)

    Javier Briones

    2011-01-01

    Full Text Available Acute Graft-versus-host disease (GVHD is a major complication after allogeneic hematopoietic stem cell transplantation. Although this process is thought to consist of several phases, T-cell activation plays a critical role in the pathogenesis of acute GVHD. To become efficient effectors, T-cells require additional costimulation after T-cell receptor signaling. A number of molecules are involved in costimulation of T-cells such as CD28, CD40L, CD30, OX40, 4-1BB, ICOS, and LIGHT. The system is regulated by inhibitory molecules, CTLA-4, and PD-1. There is experimental evidence that those molecules are implicated in the pathogenesis of GHVD. We describe how these molecules are involved in acute GVHD and how the blockade of costimulatory molecules may have potential implications for the treatment of patients with acute GVHD.

  18. Complexities in human herpesvirus-6A and -6B binding to host cells

    DEFF Research Database (Denmark)

    Pedersen, Simon Metz; Höllsberg, Per

    2006-01-01

    Human herpesvirus-6A and -6B uses the cellular receptor CD46 for fusion and infection of the host cell. The viral glycoprotein complex gH-gL from HHV-6A binds to the short consensus repeat 2 and 3 in CD46. Although all the major isoforms of CD46 bind the virus, certain isoforms may have higher...... affinity than others for the virus. Within recent years, elucidation of the viral complex has identified additional HHV-6A and -6B specific glycoproteins. Thus, gH-gL associates with a gQ1-gQ2 dimer to form a heterotetrameric complex. In addition, a novel complex consisting of gH-gL-gO has been described...... that does not bind CD46. Accumulating evidence suggests that an additional HHV-6A and -6B receptor exists. The previous simple picture of HHV-6A/B-host cell contact therefore includes more layers of complexities on both the viral and the host cell side of the interaction....

  19. Adhesion and host cell modulation: critical pathogenicity determinants of Bartonella henselae.

    Science.gov (United States)

    Franz, Bettina; Kempf, Volkhard A J

    2011-04-13

    Bartonella henselae, the agent of cat scratch disease and the vasculoproliferative disorders bacillary angiomatosis and peliosis hepatis, contains to date two groups of described pathogenicity factors: adhesins and type IV secretion systems. Bartonella adhesin A (BadA), the Trw system and possibly filamentous hemagglutinin act as promiscous or specific adhesins, whereas the virulence locus (Vir)B/VirD4 type IV secretion system modulates a variety of host cell functions. BadA mediates bacterial adherence to endothelial cells and extracellular matrix proteins and triggers the induction of angiogenic gene programming. The VirB/VirD4 type IV secretion system is responsible for, e.g., inhibition of host cell apoptosis, bacterial persistence in erythrocytes, and endothelial sprouting. The Trw-conjugation system of Bartonella spp. mediates host-specific adherence to erythrocytes. Filamentous hemagglutinins represent additional potential pathogenicity factors which are not yet characterized. The exact molecular functions of these pathogenicity factors and their contribution to an orchestral interplay need to be analyzed to understand B. henselae pathogenicity in detail.

  20. Adhesion and host cell modulation: critical pathogenicity determinants of Bartonella henselae

    Directory of Open Access Journals (Sweden)

    Kempf Volkhard AJ

    2011-04-01

    Full Text Available Abstract Bartonella henselae, the agent of cat scratch disease and the vasculoproliferative disorders bacillary angiomatosis and peliosis hepatis, contains to date two groups of described pathogenicity factors: adhesins and type IV secretion systems. Bartonella adhesin A (BadA, the Trw system and possibly filamentous hemagglutinin act as promiscous or specific adhesins, whereas the virulence locus (VirB/VirD4 type IV secretion system modulates a variety of host cell functions. BadA mediates bacterial adherence to endothelial cells and extracellular matrix proteins and triggers the induction of angiogenic gene programming. The VirB/VirD4 type IV secretion system is responsible for, e.g., inhibition of host cell apoptosis, bacterial persistence in erythrocytes, and endothelial sprouting. The Trw-conjugation system of Bartonella spp. mediates host-specific adherence to erythrocytes. Filamentous hemagglutinins represent additional potential pathogenicity factors which are not yet characterized. The exact molecular functions of these pathogenicity factors and their contribution to an orchestral interplay need to be analyzed to understand B. henselae pathogenicity in detail.

  1. Adhesion and host cell modulation: critical pathogenicity determinants of Bartonella henselae.

    Science.gov (United States)

    Franz, Bettina; Kempf, Volkhard A J

    2011-01-01

    Bartonella henselae, the agent of cat scratch disease and the vasculoproliferative disorders bacillary angiomatosis and peliosis hepatis, contains to date two groups of described pathogenicity factors: adhesins and type IV secretion systems. Bartonella adhesin A (BadA), the Trw system and possibly filamentous hemagglutinin act as promiscous or specific adhesins, whereas the virulence locus (Vir)B/VirD4 type IV secretion system modulates a variety of host cell functions. BadA mediates bacterial adherence to endothelial cells and extracellular matrix proteins and triggers the induction of angiogenic gene programming. The VirB/VirD4 type IV secretion system is responsible for, e.g., inhibition of host cell apoptosis, bacterial persistence in erythrocytes, and endothelial sprouting. The Trw-conjugation system of Bartonella spp. mediates host-specific adherence to erythrocytes. Filamentous hemagglutinins represent additional potential pathogenicity factors which are not yet characterized. The exact molecular functions of these pathogenicity factors and their contribution to an orchestral interplay need to be analyzed to understand B. henselae pathogenicity in detail. PMID:21489243

  2. Alteration of cardiac progenitor cell potency in GRMD dogs.

    Science.gov (United States)

    Cassano, M; Berardi, E; Crippa, S; Toelen, J; Barthelemy, I; Micheletti, R; Chuah, M; Vandendriessche, T; Debyser, Z; Blot, S; Sampaolesi, M

    2012-01-01

    Among the animal models of Duchenne muscular dystrophy (DMD), the Golden Retriever muscular dystrophy (GRMD) dog is considered the best model in terms of size and pathological onset of the disease. As in human patients presenting with DMD or Becker muscular dystrophies (BMD), the GRMD is related to a spontaneous X-linked mutation of dystrophin and is characterized by myocardial lesions. In this respect, GRMD is a useful model to explore cardiac pathogenesis and for the development of therapeutic protocols. To investigate whether cardiac progenitor cells (CPCs) isolated from healthy and GRMD dogs may differentiate into myocardial cell types and to test the feasibility of cell therapy for cardiomyopathies in a preclinical model of DMD, CPCs were isolated from cardiac biopsies of healthy and GRMD dogs. Gene profile analysis revealed an active cardiac transcription network in both healthy and GRMD CPCs. However, GRMD CPCs showed impaired self-renewal and cardiac differentiation. Population doubling and telomerase analyses highlighted earlier senescence and proliferation impairment in progenitors isolated from GRMD cardiac biopsies. Immunofluorescence analysis revealed that only wt CPCs showed efficient although not terminal cardiac differentiation, consistent with the upregulation of cardiac-specific proteins and microRNAs. Thus, the pathological condition adversely influences the cardiomyogenic differentiation potential of cardiac progenitors. Using PiggyBac transposon technology we marked CPCs for nuclear dsRed expression, providing a stable nonviral gene marking method for in vivo tracing of CPCs. Xenotransplantation experiments in neonatal immunodeficient mice revealed a valuable contribution of CPCs to cardiomyogenesis with homing differences between wt and dystrophic progenitors. These results suggest that cardiac degeneration in dystrophinopathies may account for the progressive exhaustion of local cardiac progenitors and shed light on cardiac stemness in

  3. Cigarette Smoke Alters the Hematopoietic Stem Cell Niche

    Directory of Open Access Journals (Sweden)

    Robert W. Siggins

    2014-02-01

    Full Text Available Effects of tobacco smoke on hematologic derangements have received little attention. This study employed a mouse model of cigarette smoke exposure to explore the effects on bone marrow niche function. While lung cancer is the most widely studied consequence of tobacco smoke exposure, other malignancies, including leukemia, are associated with tobacco smoke exposure. Animals received cigarette smoke exposure for 6 h/day, 5 days/week for 9 months. Results reveal that the hematopoietic stem and progenitor cell (HSPC pool size is reduced by cigarette smoke exposure. We next examined the effect of cigarette smoke exposure on one supporting cell type of the niche, the mesenchymal stromal cells (MSCs. Smoke exposure decreased the number of MSCs. Transplantation of naïve HSPCs into irradiated mice with cigarette smoke exposure yielded fewer numbers of engrafted HSPCs. This result suggests that smoke-exposed mice possess dysfunctional niches, resulting in abnormal hematopoiesis. Co-culture experiments using MSCs isolated from control or cigarette smoke-exposed mice with naïve HSPCs in vitro showed that MSCs from cigarette smoke-exposed mice generated marked expansion of naïve HSPCs. These data show that cigarette smoke exposure decreases in vivo MSC and HSC number and also increases pro-proliferative gene expression by cigarette smoke-exposed MSCs, which may stimulate HSPC expansion. These results of this investigation are clinically relevant to both bone marrow donors with a history of smoking and bone marrow transplant (BMT recipients with a history of smoking.

  4. The Critical Role Of VP1 In Forming The Necessary Cavities For Receptor-mediated Entry Of FMDV To The Host Cell

    OpenAIRE

    Jahanshah Ashkani; Rees, D. J. G.

    2016-01-01

    The antigenic inconsistency of the foot-and-mouth disease virus (FMDV) is very broad, such that a vaccine made from one isolate will not offer protection against infection with other isolates from the same serotype. Viral particles (VPs) or surface exposed capsid proteins, VP1–VP3, of FMDV determine both the antigenicity of the virus and its receptor-mediated entry into the host cell. Therefore, modifications of these structural proteins may alter the properties of the virus. Here we show put...

  5. PDGF induced microRNA alterations in cancer cells

    OpenAIRE

    Shao, Minghai; Rossi, Simona; Chelladurai, Bhadrani; Shimizu, Masayoshi; Ntukogu, Obiageli; Ivan, Mircea; Calin, George A.; Matei, Daniela

    2011-01-01

    Platelet derived growth factor (PDGF) regulates gene transcription by binding to specific receptors. PDGF plays a critical role in oncogenesis in brain and other tumors, regulates angiogenesis, and remodels the stroma in physiologic conditions. Here, we show by using microRNA (miR) arrays that PDGFs regulate the expression and function of miRs in glioblastoma and ovarian cancer cells. The two PDGF ligands AA and BB affect expression of several miRs in ligand-specific manner; the most robust c...

  6. A Trichomonas vaginalis Rhomboid Protease and Its Substrate Modulate Parasite Attachment and Cytolysis of Host Cells.

    Science.gov (United States)

    Riestra, Angelica M; Gandhi, Shiv; Sweredoski, Michael J; Moradian, Annie; Hess, Sonja; Urban, Sinisa; Johnson, Patricia J

    2015-12-01

    Trichomonas vaginalis is an extracellular eukaryotic parasite that causes the most common, non-viral sexually transmitted infection worldwide. Although disease burden is high, molecular mechanisms underlying T. vaginalis pathogenesis are poorly understood. Here, we identify a family of putative T. vaginalis rhomboid proteases and demonstrate catalytic activity for two, TvROM1 and TvROM3, using a heterologous cell cleavage assay. The two T. vaginalis intramembrane serine proteases display different subcellular localization and substrate specificities. TvROM1 is a cell surface membrane protein and cleaves atypical model rhomboid protease substrates, whereas TvROM3 appears to localize to the Golgi apparatus and recognizes a typical model substrate. To identify TvROM substrates, we interrogated the T. vaginalis surface proteome using both quantitative proteomic and bioinformatic approaches. Of the nine candidates identified, TVAG_166850 and TVAG_280090 were shown to be cleaved by TvROM1. Comparison of amino acid residues surrounding the predicted cleavage sites of TvROM1 substrates revealed a preference for small amino acids in the predicted transmembrane domain. Over-expression of TvROM1 increased attachment to and cytolysis of host ectocervical cells. Similarly, mutations that block the cleavage of a TvROM1 substrate lead to its accumulation on the cell surface and increased parasite adherence to host cells. Together, these data indicate a role for TvROM1 and its substrate(s) in modulating attachment to and lysis of host cells, which are key processes in T. vaginalis pathogenesis.

  7. The Commensal Streptococcus salivarius K12 Downregulates the Innate Immune Responses of Human Epithelial Cells and Promotes Host-Microbe Homeostasis▿ †

    Science.gov (United States)

    Cosseau, Celine; Devine, Deirdre A.; Dullaghan, Edie; Gardy, Jennifer L.; Chikatamarla, Avinash; Gellatly, Shaan; Yu, Lorraine L.; Pistolic, Jelena; Falsafi, Reza; Tagg, John; Hancock, Robert E. W.

    2008-01-01

    Streptococcus salivarius is an early colonizer of human oral and nasopharyngeal epithelia, and strain K12 has reported probiotic effects. An emerging paradigm indicates that commensal bacteria downregulate immune responses through the action on NF-κB signaling pathways, but additional mechanisms underlying probiotic actions are not well understood. Our objective here was to identify host genes specifically targeted by K12 by comparing their responses with responses elicited by pathogens and to determine if S. salivarius modulates epithelial cell immune responses. RNA was extracted from human bronchial epithelial cells (16HBE14O- cells) cocultured with K12 or bacterial pathogens. cDNA was hybridized to a human 21K oligonucleotide-based array. Data were analyzed using ArrayPipe, InnateDB, PANTHER, and oPOSSUM. Interleukin 8 (IL-8) and growth-regulated oncogene alpha (Groα) secretion were determined by enzyme-linked immunosorbent assay. It was demonstrated that S. salivarius K12 specifically altered the expression of 565 host genes, particularly those involved in multiple innate defense pathways, general epithelial cell function and homeostasis, cytoskeletal remodeling, cell development and migration, and signaling pathways. It inhibited baseline IL-8 secretion and IL-8 responses to LL-37, Pseudomonas aeruginosa, and flagellin in epithelial cells and attenuated Groα secretion in response to flagellin. Immunosuppression was coincident with the inhibition of activation of the NF-κB pathway. Thus, the commensal and probiotic behaviors of S. salivarius K12 are proposed to be due to the organism (i) eliciting no proinflammatory response, (ii) stimulating an anti-inflammatory response, and (iii) modulating genes associated with adhesion to the epithelial layer and homeostasis. S. salivarius K12 might thereby ensure that it is tolerated by the host and maintained on the epithelial surface while actively protecting the host from inflammation and apoptosis induced by

  8. The commensal Streptococcus salivarius K12 downregulates the innate immune responses of human epithelial cells and promotes host-microbe homeostasis.

    Science.gov (United States)

    Cosseau, Celine; Devine, Deirdre A; Dullaghan, Edie; Gardy, Jennifer L; Chikatamarla, Avinash; Gellatly, Shaan; Yu, Lorraine L; Pistolic, Jelena; Falsafi, Reza; Tagg, John; Hancock, Robert E W

    2008-09-01

    Streptococcus salivarius is an early colonizer of human oral and nasopharyngeal epithelia, and strain K12 has reported probiotic effects. An emerging paradigm indicates that commensal bacteria downregulate immune responses through the action on NF-kappaB signaling pathways, but additional mechanisms underlying probiotic actions are not well understood. Our objective here was to identify host genes specifically targeted by K12 by comparing their responses with responses elicited by pathogens and to determine if S. salivarius modulates epithelial cell immune responses. RNA was extracted from human bronchial epithelial cells (16HBE14O- cells) cocultured with K12 or bacterial pathogens. cDNA was hybridized to a human 21K oligonucleotide-based array. Data were analyzed using ArrayPipe, InnateDB, PANTHER, and oPOSSUM. Interleukin 8 (IL-8) and growth-regulated oncogene alpha (Groalpha) secretion were determined by enzyme-linked immunosorbent assay. It was demonstrated that S. salivarius K12 specifically altered the expression of 565 host genes, particularly those involved in multiple innate defense pathways, general epithelial cell function and homeostasis, cytoskeletal remodeling, cell development and migration, and signaling pathways. It inhibited baseline IL-8 secretion and IL-8 responses to LL-37, Pseudomonas aeruginosa, and flagellin in epithelial cells and attenuated Groalpha secretion in response to flagellin. Immunosuppression was coincident with the inhibition of activation of the NF-kappaB pathway. Thus, the commensal and probiotic behaviors of S. salivarius K12 are proposed to be due to the organism (i) eliciting no proinflammatory response, (ii) stimulating an anti-inflammatory response, and (iii) modulating genes associated with adhesion to the epithelial layer and homeostasis. S. salivarius K12 might thereby ensure that it is tolerated by the host and maintained on the epithelial surface while actively protecting the host from inflammation and apoptosis

  9. Radiation-induced alterations of histone post-translational modification levels in lymphoblastoid cell lines

    International Nuclear Information System (INIS)

    Radiation-induced alterations in posttranslational histone modifications (PTMs) may affect the cellular response to radiation damage in the DNA. If not reverted appropriately, altered PTM patterns may cause long-term alterations in gene expression regulation and thus lead to cancer. It is therefore important to characterize radiation-induced alterations in PTM patterns and the factors affecting them. A lymphoblastoid cell line established from a normal donor was used to screen for alterations in methylation levels at H3K4, H3K9, H3K27, and H4K20, as well as acetylation at H3K9, H3K56, H4K5, and H4K16, by quantitative Western Blot analysis at 15 min, 1 h and 24 h after irradiation with 2 Gy and 10 Gy. The variability of alterations in acetylation marks was in addition investigated in a panel of lymphoblastoid cell lines with differing radiosensitivity established from lung cancer patients. The screening procedure demonstrated consistent hypomethylation at H3K4me3 and hypoacetylation at all acetylation marks tested. In the panel of lymphoblastoid cell lines, however, a high degree of inter-individual variability became apparent. Radiosensitive cell lines showed more pronounced and longer lasting H4K16 hypoacetylation than radioresistant lines, which correlates with higher levels of residual γ-H2AX foci after 24 h. So far, the factors affecting extent and duration of radiation-induced histone alterations are poorly defined. The present work hints at a high degree of inter-individual variability and a potential correlation of DNA damage repair capacity and alterations in PTM levels

  10. Gene expression profiling in host-pathogen interactions and identification of the molecular mechanisms involved in dendrictic cells activation

    OpenAIRE

    Torri,, M.

    2009-01-01

    In this thesis we used a functional genomic approach to study host-pathogen interactions [1]. We analyzed the interaction from the host point of view and in particular from the dendritic cells point of view. Dendritic cells (DCs) constitute a heterogeneous group of antigen-presenting leukocytes important in activation of both innate and adaptive immunity [2]. In the first part of this thesis we explored the possibility to use dendritic cell transcriptomes to generate biomarkers of inflamma...

  11. Prenatal cadmium exposure alters postnatal immune cell development and function

    Energy Technology Data Exchange (ETDEWEB)

    Hanson, Miranda L.; Holásková, Ida; Elliott, Meenal; Brundage, Kathleen M.; Schafer, Rosana; Barnett, John B., E-mail: jbarnett@hsc.wvu.edu

    2012-06-01

    Cadmium (Cd) is generally found in low concentrations in the environment due to its widespread and continual use, however, its concentration in some foods and cigarette smoke is high. Although evidence demonstrates that adult exposure to Cd causes changes in the immune system, there are limited reports of immunomodulatory effects of prenatal exposure to Cd. This study was designed to investigate the effects of prenatal exposure to Cd on the immune system of the offspring. Pregnant C57Bl/6 mice were exposed to an environmentally relevant dose of CdCl{sub 2} (10 ppm) and the effects on the immune system of the offspring were assessed at two time points following birth (2 and 7 weeks of age). Thymocyte and splenocyte phenotypes were analyzed by flow cytometry. Prenatal Cd exposure did not affect thymocyte populations at 2 and 7 weeks of age. In the spleen, the only significant effect on phenotype was a decrease in the number of macrophages in male offspring at both time points. Analysis of cytokine production by stimulated splenocytes demonstrated that prenatal Cd exposure decreased IL-2 and IL-4 production by cells from female offspring at 2 weeks of age. At 7 weeks of age, splenocyte IL-2 production was decreased in Cd-exposed males while IFN-γ production was decreased from both male and female Cd-exposed offspring. The ability of the Cd-exposed offspring to respond to immunization with a S. pneumoniae vaccine expressing T-dependent and T-independent streptococcal antigens showed marked increases in the levels of both T-dependent and T-independent serum antibody levels compared to control animals. CD4{sup +}FoxP3{sup +}CD25{sup +} (nTreg) cell percentages were increased in the spleen and thymus in all Cd-exposed offspring except in the female spleen where a decrease was seen. CD8{sup +}CD223{sup +} T cells were markedly decreased in the spleens in all offspring at 7 weeks of age. These findings suggest that even very low levels of Cd exposure during gestation can

  12. Interactions between mycoplasma lipid-associated membrane proteins and the host cells

    Institute of Scientific and Technical Information of China (English)

    YOU Xiao-xing; ZENG Yan-hua; WU Yi-mou

    2006-01-01

    Mycoplamas are a group of wall-less prokaryotes widely distributed in nature, some of which are pathogenic for humans and animals. There are many lipoproteins anchored on the outer face of the plasma membrane, called lipid-associated membrane proteins (LAMPs). LAMPs are highly antigenic and could undergo phase and size variation, and are recognized by the innate immune system through Toll-like receptors (TLR) 2 and 6. LAMPs can modulate the immune system, and could induce immune cells apoptosis or death. In addition, they may associate with malignant transformation of host cells and are also considered to be cofactors in the progression of AIDS.

  13. Sialoglycoconjugates in Trypanosoma cruzi-host cell interaction: possible biological model - a review

    Directory of Open Access Journals (Sweden)

    Alane Beatriz Vermelho

    1994-03-01

    Full Text Available A number of glycoconjugates, including glycolipids and glycoproteins, participate in the process of host-cell invasion by Trypanosoma cruzi and one of the most important carbohydrates involved on this interaction is sialic acid. It is known that parasite trans-sialidase participates with sialic acid in a coordinated fashion in the initial stages of invasion. Given the importance of these sialogycoconjugates, this review sets out various possible biological models for the interaction between the parasite and mammalian cells that possess a sialylated receptor/ligand system.

  14. Discovery and targeted LC-MS/MS of purified polerovirus reveals differences in the virus-host interactome associated with altered aphid transmission.

    Directory of Open Access Journals (Sweden)

    Michelle Cilia

    Full Text Available Circulative transmission of viruses in the Luteoviridae, such as cereal yellow dwarf virus (CYDV, requires a series of precisely orchestrated interactions between virus, plant, and aphid proteins. Natural selection has favored these viruses to be retained in the phloem to facilitate acquisition and transmission by aphids. We show that treatment of infected oat tissue homogenate with sodium sulfite reduces transmission of the purified virus by aphids. Transmission electron microscopy data indicated no gross change in virion morphology due to treatments. However, treated virions were not acquired by aphids through the hindgut epithelial cells and were not transmitted when injected directly into the hemocoel. Analysis of virus preparations using nanoflow liquid chromatography coupled to tandem mass spectrometry revealed a number of host plant proteins co-purifying with viruses, some of which were lost following sodium sulfite treatment. Using targeted mass spectrometry, we show data suggesting that several of the virus-associated host plant proteins accumulated to higher levels in aphids that were fed on CYDV-infected plants compared to healthy plants. We propose two hypotheses to explain these observations, and these are not mutually exclusive: (a that sodium sulfite treatment disrupts critical virion-host protein interactions required for aphid transmission, or (b that host infection with CYDV modulates phloem protein expression in a way that is favorable for virus uptake by aphids. Importantly, the genes coding for the plant proteins associated with virus may be examined as targets in breeding cereal crops for new modes of virus resistance that disrupt phloem-virus or aphid-virus interactions.

  15. Discovery and targeted LC-MS/MS of purified polerovirus reveals differences in the virus-host interactome associated with altered aphid transmission.

    Science.gov (United States)

    Cilia, Michelle; Peter, Kari A; Bereman, Michael S; Howe, Kevin; Fish, Tara; Smith, Dawn; Gildow, Fredrick; MacCoss, Michael J; Thannhauser, Theodore W; Gray, Stewart M

    2012-01-01

    Circulative transmission of viruses in the Luteoviridae, such as cereal yellow dwarf virus (CYDV), requires a series of precisely orchestrated interactions between virus, plant, and aphid proteins. Natural selection has favored these viruses to be retained in the phloem to facilitate acquisition and transmission by aphids. We show that treatment of infected oat tissue homogenate with sodium sulfite reduces transmission of the purified virus by aphids. Transmission electron microscopy data indicated no gross change in virion morphology due to treatments. However, treated virions were not acquired by aphids through the hindgut epithelial cells and were not transmitted when injected directly into the hemocoel. Analysis of virus preparations using nanoflow liquid chromatography coupled to tandem mass spectrometry revealed a number of host plant proteins co-purifying with viruses, some of which were lost following sodium sulfite treatment. Using targeted mass spectrometry, we show data suggesting that several of the virus-associated host plant proteins accumulated to higher levels in aphids that were fed on CYDV-infected plants compared to healthy plants. We propose two hypotheses to explain these observations, and these are not mutually exclusive: (a) that sodium sulfite treatment disrupts critical virion-host protein interactions required for aphid transmission, or (b) that host infection with CYDV modulates phloem protein expression in a way that is favorable for virus uptake by aphids. Importantly, the genes coding for the plant proteins associated with virus may be examined as targets in breeding cereal crops for new modes of virus resistance that disrupt phloem-virus or aphid-virus interactions. PMID:23118947

  16. Endothelial Cell Morphology and Migration are Altered by Changes in Gravitational Fields

    Science.gov (United States)

    Melhado, Caroline; Sanford, Gary; Harris-Hooker, Sandra

    1997-01-01

    Many of the physiological changes of the cardiovascular system during space flight may originate from the dysfunction of basic biological mechanisms caused by microgravity. The weightlessness affects the system when blood and other fluids move to the upper body causing the heart to enlarge to handle the increased blood flow to the upper extremities and decrease circulating volume. Increase arterial pressure triggers baroreceptors which signal the brain to adjust heart rate. Hemodynarnic studies indicate that the microgravity-induced headward fluid redistribution results in various cardiovascular changes such as; alteration of vascular permeability resulting in lipid accumulation in the lumen of the vasculature and degeneration of the the vascular wall, capillary alteration with extensive endothelial invagination. Achieving a true microgravity environment in ground based studies for prolonged periods is virtually impossible. The application of vector-averaged gravity to mammalian cells using horizontal clinostat produces alterations of cellular behavior similar to those observed in microgravity. Similarly, the low shear, horizontally rotating bioreactor (originally designed by NASA) also duplicates several properties of microgravity. Additionally, increasing gravity, i.e., hypcrgravity is easily achieved. Hypergravity has been found to increase the proliferation of several different cell lines (e.g., chick embryo fibroblasts) while decreasing cell motility and slowing liver regeneration following partial hepatectomy. The effect of altered gravity on cells maybe similar to those of other physical forces, i.e. shear stress. Previous studies examining laminar flow and shear stress on endothelial cells found that the cells elongate, orient with the direction of flow, and reorganize their F-actin structure, with concomitant increase in cell stiffness. These studies suggest that alterations in the gravity environment will change the behavior of most cells, including

  17. IL4RA on lymphatic endothelial cells promotes T cell egress during sclerodermatous graft versus host disease

    Science.gov (United States)

    Urso, Katia; Alvarez, David; Cremasco, Viviana; Tsang, Kelly; Grauel, Angelo; Lafyatis, Robert; von Andrian, Ulrich H.; Ermann, Joerg; Aliprantis, Antonios O.

    2016-01-01

    Systemic sclerosis (SSc) is a potentially fatal autoimmune disorder with limited therapeutic options. Sclerodermatous graft versus host disease (sclGvHD), induced by transfer of B10.D2 splenocytes into BALB/c Rag2−/− mice, models an inflammatory subset of SSc characterized by a prominent IL13-induced gene expression signature in the skin. Host mice deficient in IL4RA, a subunit of the type II IL4/IL13 receptor, are protected from sclGvHD. While IL4RA has a well-established role in Th2 differentiation and alternative macrophage activation, we report here a previously unappreciated function for IL4RA in lymphatic endothelial cells (LECs): regulation of activated T cell egress. Seven days after splenocyte transfer, Il4ra−/− hosts had increased numbers of activated graft CD4+ T cells in skin draining lymph nodes (dLNs) but fewer T cells in efferent lymph, blood, and skin. Sphingosine-1 phosphate (S1P), master regulator of lymphocyte egress from LNs, was lower in dLNs of Il4ra−/− hosts with a corresponding decrease of S1P kinase 1 (Sphk1) expression in LECs. Bypassing the efferent lymphatics via i.v. injection of CD4+ T cells from dLNs of Il4ra−/− sclGvHD mice restored clinical GvHD in secondary Il4ra−/− recipients. These results identify a role for IL4RA and suggest that modulation of lymphocyte egress from LNs may be effective in SSc and GvHD.

  18. Confocal Fluorescence Imaging Enables Noninvasive Quantitative Assessment of Host Cell Populations In Vivo Following Photodynamic Therapy

    Directory of Open Access Journals (Sweden)

    Soumya Mitra, Oleg Mironov, Thomas H. Foster

    2012-01-01

    Full Text Available We report the use of optical imaging strategies to noninvasively examine photosensitizer distribution and physiological and host responses to 2-[1-hexyloxyethyl]-2 devinyl pyropheophorbide-a (HPPH-mediated photodynamic therapy (PDT of EMT6 tumors established in the ears of BALB/c mice. 24 h following intravenous (IV administration of 1 μmol kg-1 HPPH, wide-field fluorescence imaging reveals tumor selectivity with an approximately 2-3-fold differential between tumor and adjacent normal tissue. Confocal microscopy demonstrates a relatively homogeneous intratumor HPPH distribution. Labeling of host cells using fluorophore-conjugated antibodies allowed the visualization of Gr1+/CD11b+ leukocytes and major histocompatibility complex class II (MHC-II+ cells in vivo. Imaging of the treated site at different time-points following irradiation shows significant and rapid increases in Gr1+ cells in response to therapy. The maximum accumulation of Gr1+ cells is found at 24 h post-irradiation, followed by a decrease at the 48 h time-point. Using IV-injected FITC-conjugated dextran as a fluorescent perfusion marker, we imaged tissue perfusion at different times post-irradiation and found that the reduced Gr1+ cell density at 48 h correlated strongly with functional damage to the vasculature as reported via decreased perfusion status. Dual color confocal imaging experiments demonstrates that about 90% of the anti-Gr1 cell population co-localized with anti-CD11b labeling, thus indicating that majority of the Gr1-labeled cells were neutrophils. At 24 h post-PDT, an approximately 2-fold increase in MHC-II+ cells relative to untreated control is also observed. Co-localization analysis reveals an increase in the fraction of Gr1+ cells expressing MHC-II, suggesting that HPPH-PDT is stimulating neutrophils to express an antigen-presenting phenotype.

  19. Host cell reactivation by human cells of DNA expression vectors damaged by ultraviolet radiation or by acid-heat treatment

    International Nuclear Information System (INIS)

    We utilized a plasmid vector host cell reactivation assay to probe the biological functioning of DNA expression vectors and their encoded genes. We studied the effect of ultraviolet radiation or acid-heat treatment on the transient expression of genes transfected into normal human cells and into DNA repair deficient (xeroderma pigmentosum) cells and modification of gene expression by sodium butyrate. The results showed that u.v. damage of DNA expression vectors was subject to repair by the normal host cells, but acid-heat treatment resulted in damage (apurinic sites) that was handled in a similar manner by excision repair deficient and excision repair proficient human cells. In both normal and xeroderma pigmentosum cells sodium butyrate treatment of cells resulted in a greater stimulation of chloramphenicol acetyltransferase expression with u.v. damaged than with undamaged plasmid. This assay thus permits examination of the effects of defined types of DNA damage on plasmid expression and study of its modulation by cellular repair activities. (author)

  20. Antimetastatic function of concomitant antitumor immunity. I. Host Ly-1+2+ effector T cells prevent the enumeration of metastatic tumor cells in a biological assay

    International Nuclear Information System (INIS)

    A mouse survival assay was evaluated for its suitability to enumerate metastatic P815 tumor cells in the draining lymph node and spleen of a B6D2 F1 (H-2/sup b/ x H-2/sup d/) host bearing a primary intradermal P815 tumor. The mouse survival assay is based on the linear relationship between the log10 number of P815 tumor cells (H-2/sup d/) injected i.p. into mice and their mean survival time. It was found that the assay is capable of quantifying as few as 10 tumor cells in lymph node and spleen, but only if cell suspensions of these organs are treated with anti-H-2/sup b/ serum and complement, in order to selectively destroy H-2/sup bd/ host cells. This was necessary because host cells from the lymph node and spleen of a tumor-bearing host possessed antitumor functions, in that they were capable of destroying the H-2/sup d/ P815 tumor cells when admixed with the tumor cells and injected i.p. into 800-rad irradiated test recipients. The kinetics of acquisition and loss of host cells with antitumor function and the Ly phenotype of these host cells suggest that they are the same cells that give the tumor-bearing host the capacity to express concomitant immunity against a tumor implant

  1. Role of Fc Gamma Receptors in Triggering Host Cell Activation and Cytokine Release by Borrelia burgdorferi

    Science.gov (United States)

    Talkington, Jeffrey; Nickell, Steven P.

    2001-01-01

    Borrelia burgdorferi, the spirochetal bacterium that causes human Lyme disease, encodes numerous lipoproteins which have the capacity to trigger the release of proinflammatory cytokines from a variety of host cell types, and it is generally believed that these cytokines contribute to the disease process in vivo. We previously reported that low-passage-number infectious B. burgdorferi spirochetes express a novel lipidation-independent activity which induces secretion of the proinflammatory cytokine tumor necrosis factor alpha (TNF-α) by the mouse MC/9 mast cell line. Using RNase protection assays, we determined that mast cells exposed in vitro to low-passage-number, but not high-passage-number, B. burgdorferi spirochetes show increased expression of additional mRNAs representing several chemokines, including macrophage-inflammatory protein 1α (MIP-1α), MIP-1β, and TCA3, as well as the proinflammatory cytokine interleukin-6. Furthermore, mast cell TNF-α secretion can be inhibited by the phosphatidylinositol 3-kinase inhibitor wortmannin and also by preincubation with purified mouse immunoglobulin G1 (IgG1) and IgG2a, but not mouse IgG3, and by a mouse Fc gamma receptor II and III (FcγRII/III)-specific rat monoclonal antibody, suggesting the likely involvement of host FcγRIII in B. burgdorferi-mediated signaling. A role for passively adsorbed rabbit or bovine IgG or serum components in B. burgdorferi-mediated FcγR signaling was excluded in control experiments. These studies confirm that low-passage-number B. burgdorferi spirochetes express a novel activity which upregulates the expression of a variety of host cell chemokine and cytokine genes, and they also establish a novel antibody-independent role for FcγRs in transduction of activation signals by bacterial products. PMID:11119532

  2. Comparative phosphoproteomics reveals components of host cell invasion and post-transcriptional regulation during Francisella infection

    Energy Technology Data Exchange (ETDEWEB)

    Nakayasu, Ernesto S.; Tempel, Rebecca; Cambronne, Xiaolu A.; Petyuk, Vladislav A.; Jones, Marcus B.; Gritsenko, Marina A.; Monroe, Matthew E.; Yang, Feng; Smith, Richard D.; Adkins, Joshua N.; Heffron, Fred

    2013-09-22

    Francisella tularensis is a facultative intracellular bacterium that causes the deadly disease tularemia. Most evidence suggests that Francisella is not well recognized by the innate immune system that normally leads to cytokine expression and cell death. In previous work, we identified new bacterial factors that were hyper-cytotoxic to macrophages. Four of the identified hyper-cytotoxic strains (lpcC, manB, manC and kdtA) had an impaired lipopolysaccharide (LPS) synthesis and produced an exposed lipid A lacking the O-antigen. These mutants were not only hyper-cytotoxic but also were phagocytosed at much higher rates compared to the wild type parent strain. To elucidate the cellular signaling underlying this enhanced phagocytosis and cell death, we performed a large-scale comparative phosphoproteomic analysis of cells infected with wild-type and delta-lpcC F. novicida. Our data suggest that not only actin but also intermediate filaments and microtubules are important for F. novicida entry into the host cells. In addition, we observed differential phosphorylation of tristetraprolin (TTP), a key component of the mRNA-degrading machinery that controls the expression of a variety of genes including many cytokines. Infection with the delta-lpcC mutant induced the hyper-phosphorylation and inhibition of TTP, leading to the production of cytokines such as IL-1beta and TNF-alpha which may kill the host cells by triggering apoptosis. Together, our data provide new insights for Francisella invasion and a post-transcriptional mechanism that prevents the expression of host immune response factors that controls infection by this pathogen.

  3. Cell wall glycoproteins at interaction sites between parasitic giant dodder (Cuscuta reflexa) and its host Pelargonium zonale.

    Science.gov (United States)

    Striberny, Bernd; Krause, Kirsten

    2015-01-01

    The process of host plant penetration by parasitic dodder (genus Cuscuta) is accompanied by molecular and structural changes at the host/parasite interface. Recently, changes in pectin methyl esterification levels in the host cell walls abutting parasitic cells in established infection sites were reported. In addition to that, we show here that the composition of cell wall glycoproteins in Cuscuta-infected Pelargonium zonale undergoes substantial changes. While several arabinogalactan protein epitopes exhibit decreased abundances in the vicinity of the Cuscuta reflexa haustorium, extensins tend to increase in the infected areas. PMID:26367804

  4. Cell wall glycoproteins at interaction sites between parasitic giant dodder (Cuscuta reflexa) and its host Pelargonium zonale.

    Science.gov (United States)

    Striberny, Bernd; Krause, Kirsten

    2015-01-01

    The process of host plant penetration by parasitic dodder (genus Cuscuta) is accompanied by molecular and structural changes at the host/parasite interface. Recently, changes in pectin methyl esterification levels in the host cell walls abutting parasitic cells in established infection sites were reported. In addition to that, we show here that the composition of cell wall glycoproteins in Cuscuta-infected Pelargonium zonale undergoes substantial changes. While several arabinogalactan protein epitopes exhibit decreased abundances in the vicinity of the Cuscuta reflexa haustorium, extensins tend to increase in the infected areas.

  5. Surface Molecules Released by Trypanosoma cruzi Metacyclic Forms Downregulate Host Cell Invasion

    Science.gov (United States)

    Clemente, Tatiana Mordente; Cortez, Cristian; Novaes, Antônio da Silva; Yoshida, Nobuko

    2016-01-01

    Background The question whether metacylic trypomastigote (MT) forms of different T. cruzi strains differentially release surface molecules, and how they affect host cell invasion, remains to be fully clarified. We addressed that question using T. cruzi strains that differ widely in the ability to invade cells. Methodology/Principal Findings Metacyclic forms were incubated at 37°C for 1 h in complete D10 medium or in nutrient-deprived PBS containing Ca2+ and Mg2+ (PBS++). The conditioned medium (CM), collected after parasite centrifugation, was used for cell invasion assays and Western blot analysis, using monoclonal antibodies directed to gp82 and gp90, the MT surface molecules that promote and negatively regulate invasion, respectively. CM of poorly invasive G strain (G-CM) contained high amounts of gp90 and gp82, either in vesicles or as soluble molecules. CM of highly invasive CL strain (CL-CM) contained gp90 and gp82 at very low levels. HeLa cells were incubated for 1 h with CL strain MT in D10, in absence or in the presence of G-CM or CL-CM. Parasite invasion was significantly inhibited by G-CM, but not by CL-CM. As G strain MT invasion rate in D10 is very low, assays with this strain were performed in PBS++, which induces invasion-promoting lysosome-spreading. G-CM, but not CL-CM, significantly inhibited G strain internalization, effect that was counteracted by preincubating G-CM with an anti-gp90 monoclonal antibody or anti-gp82 polyclonal antibody that do not recognize live MT. G strain CM generated in PBS++ contained much lower amounts of gp90 and gp82 as compared to CM produced in D10, and exhibited lower inhibitory effect on host cell invasion. Conclusion/Significance Our data suggest that the surface molecules spontaneously released by MT impair parasite-host cell interaction, gp82 presumably competing with the molecule expressed on MT surface for the host cell receptor, and gp90 further contributing to down modulate invasion. PMID:27483135

  6. Essential domains of Anaplasma phagocytophilum invasins utilized to infect mammalian host cells.

    Directory of Open Access Journals (Sweden)

    David Seidman

    2015-02-01

    Full Text Available Anaplasma phagocytophilum causes granulocytic anaplasmosis, an emerging disease of humans and domestic animals. The obligate intracellular bacterium uses its invasins OmpA, Asp14, and AipA to infect myeloid and non-phagocytic cells. Identifying the domains of these proteins that mediate binding and entry, and determining the molecular basis of their interactions with host cell receptors would significantly advance understanding of A. phagocytophilum infection. Here, we identified the OmpA binding domain as residues 59 to 74. Polyclonal antibody generated against a peptide spanning OmpA residues 59 to 74 inhibited A. phagocytophilum infection of host cells and binding to its receptor, sialyl Lewis x (sLe(x-capped P-selectin glycoprotein ligand 1. Molecular docking analyses predicted that OmpA residues G61 and K64 interact with the two sLe(x sugars that are important for infection, α2,3-sialic acid and α1,3-fucose. Amino acid substitution analyses demonstrated that K64 was necessary, and G61 was contributory, for recombinant OmpA to bind to host cells and competitively inhibit A. phagocytophilum infection. Adherence of OmpA to RF/6A endothelial cells, which express little to no sLe(x but express the structurally similar glycan, 6-sulfo-sLe(x, required α2,3-sialic acid and α1,3-fucose and was antagonized by 6-sulfo-sLe(x antibody. Binding and uptake of OmpA-coated latex beads by myeloid cells was sensitive to sialidase, fucosidase, and sLe(x antibody. The Asp14 binding domain was also defined, as antibody specific for residues 113 to 124 inhibited infection. Because OmpA, Asp14, and AipA each contribute to the infection process, it was rationalized that the most effective blocking approach would target all three. An antibody cocktail targeting the OmpA, Asp14, and AipA binding domains neutralized A. phagocytophilum binding and infection of host cells. This study dissects OmpA-receptor interactions and demonstrates the effectiveness of binding

  7. Alterations of FHIT Gene and P16 Gene in Nickel Transformed Human Bronchial Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    WEI-DONG JI; JIA-KUN CHEN; JIA-CHUN LU; ZHONG-LIANG WU; FEI YI; SU-MEI FENG

    2006-01-01

    To study the alterations of FHIT gene and P16 gene in malignant transformed human bronchial epithelial cells induced by crystalline nickel sulfide using an immoral human bronchial epithelial cell line, and to explore the molecular mechanism of nickel carcinogenesis. Methods 16HBE cells were treated 6 times with different concentrations of NiS in vitro, and the degree of malignant transformation was determined by assaying the anchorage-independent growth and tumorigenicity. Malignant transformed cells and tumorigenic cells were examined for alterations of FHIT gene and P16 gene using RT-PCR, DNA sequencing, silver staining PCR-SSCP and Western blotting. Results NiS-treated cells exhibited overlapping growth. Compared with that of negative control cells, soft agar colony formation efficiency of NiS-treated cells showed significant increases (P<0.01) and dose-dependent effects. NiS-treated cells could form tumors in nude mice, and a squamous cell carcinoma was confirmed by histopathological examination. No mutation of exon 2 and exons 2-3, no abnormal expression in p16 gene and mutation of FHIT exons 5-8 and exons 1-4 or exons 5-9 were observed in transformed cells and tumorigenic cells. However, aberrant transcripts or loss of expression of the FHIT gene and Fhit protein was observed in transformed cells and tumorigenic cells. One of the aberrant transcripts in the FHIT gene was confirmed to have a deletion of exon 6, exon 7, exon 8, and an insertion of a 36 bp sequence replacing exon 6-8. Conclusions The FHIT gene rather than the P16 gene, plays a definite role in nickel carcinogenesis. Alterations of the FHIT gene induced by crystalline NiS may be a molecular event associated with carcinogen, chromosome fragile site instability and cell malignant transformation. FHIT may be an important target gene activated by nickel and other exotic carcinogens.

  8. Altered Cell Mechanics from the Inside: Dispersed Single Wall Carbon Nanotubes Integrate with and Restructure Actin

    Directory of Open Access Journals (Sweden)

    Mohammad F. Islam

    2012-05-01

    Full Text Available With a range of desirable mechanical and optical properties, single wall carbon nanotubes (SWCNTs are a promising material for nanobiotechnologies. SWCNTs also have potential as biomaterials for modulation of cellular structures. Previously, we showed that highly purified, dispersed SWCNTs grossly alter F-actin inside cells. F-actin plays critical roles in the maintenance of cell structure, force transduction, transport and cytokinesis. Thus, quantification of SWCNT-actin interactions ranging from molecular, sub-cellular and cellular levels with both structure and function is critical for developing SWCNT-based biotechnologies. Further, this interaction can be exploited, using SWCNTs as a unique actin-altering material. Here, we utilized molecular dynamics simulations to explore the interactions of SWCNTs with actin filaments. Fluorescence lifetime imaging microscopy confirmed that SWCNTs were located within ~5 nm of F-actin in cells but did not interact with G-actin. SWCNTs did not alter myosin II sub-cellular localization, and SWCNT treatment in cells led to significantly shorter actin filaments. Functionally, cells with internalized SWCNTs had greatly reduced cell traction force. Combined, these results demonstrate direct, specific SWCNT alteration of F-actin structures which can be exploited for SWCNT-based biotechnologies and utilized as a new method to probe fundamental actin-related cellular processes and biophysics.

  9. Genomic Modifiers of Natural Killer Cells, Immune Responsiveness and Lymphoid Tissue Remodeling Together Increase Host Resistance to Viral Infection.

    Science.gov (United States)

    Gillespie, Alyssa Lundgren; Teoh, Jeffrey; Lee, Heather; Prince, Jessica; Stadnisky, Michael D; Anderson, Monique; Nash, William; Rival, Claudia; Wei, Hairong; Gamache, Awndre; Farber, Charles R; Tung, Kenneth; Brown, Michael G

    2016-02-01

    The MHC class I D(k) molecule supplies vital host resistance during murine cytomegalovirus (MCMV) infection. Natural killer (NK) cells expressing the Ly49G2 inhibitory receptor, which specifically binds D(k), are required to control viral spread. The extent of D(k)-dependent host resistance, however, differs significantly amongst related strains of mice, C57L and MA/My. As a result, we predicted that relatively small-effect modifier genetic loci might together shape immune cell features, NK cell reactivity, and the host immune response to MCMV. A robust D(k)-dependent genetic effect, however, has so far hindered attempts to identify additional host resistance factors. Thus, we applied genomic mapping strategies and multicolor flow cytometric analysis of immune cells in naive and virus-infected hosts to identify genetic modifiers of the host immune response to MCMV. We discovered and validated many quantitative trait loci (QTL); these were mapped to at least 19 positions on 16 chromosomes. Intriguingly, one newly discovered non-MHC locus (Cmv5) controlled splenic NK cell accrual, secondary lymphoid organ structure, and lymphoid follicle development during MCMV infection. We infer that Cmv5 aids host resistance to MCMV infection by expanding NK cells needed to preserve and protect essential tissue structural elements, to enhance lymphoid remodeling and to increase viral clearance in spleen.

  10. Early shutoff of host protein synthesis in cells infected with herpes simplex viruses.

    Science.gov (United States)

    Matis, J; Kúdelová, M

    2001-01-01

    Herpes simplex viruses 1 (HSV-1) and 2 (HSV-2) are capable of suppressing the host cell protein synthesis even without viral gene expression. This phenomenon is known as the early shutoff or as the virion-associated host shutoff (vhs) to emphasize that it is mediated by a component of infecting virions which is a product of the UL41 (vhs) gene. The UL41 encoded protein is a functional tegument protein also present in light (L) particles and is not essential for virus replication. The major product of UL41 gene is a 58 K phosphoprotein. At least two forms of UL41 protein differing in the extent of phosphorylation are present in HSV-1-infected cells. HSV-2 compared to HSV-1 strains display a stronger vhs phenotype. However, in superinfection experiments the less strong vhs phenotype is dominant. UL41 protein triggers disruption of polysomes and rapid degradation of all host and viral mRNAs and blocks a reporter gene expression without other HSVs proteins. The available evidence suggests that UL41 protein is either itself a ribonuclease (RNase) or a subunit of RNase that contains also one or more cellular subunits. UL41 protein is capable of interacting with a transactivator of an alpha-gene, the alpha-transinducing factor (alpha-TIF). Interaction of UL41 protein with alpha-TIF down regulates the UL41 (vhs) gene activity during lytic infection. The possible role of other viral proteins in the shutoff is discussed.

  11. Do oral bacteria alter the regenerative potential of stem cells? A concise review.

    Science.gov (United States)

    Chatzivasileiou, Kyriaki; Kriebel, Katja; Steinhoff, Gustav; Kreikemeyer, Bernd; Lang, Hermann

    2015-09-01

    Mesenchymal stem cells (MSCs) are widely recognized as critical players in tissue regeneration. New insights into stem cell biology provide evidence that MSCs may also contribute to host defence and inflammation. In case of tissue injury or inflammatory diseases, e.g. periodontitis, stem cells are mobilized towards the site of damage, thus coming in close proximity to bacteria and bacterial components. Specifically, in the oral cavity, complex ecosystems of commensal bacteria live in a mutually beneficial state with the host. However, the formation of polymicrobial biofilm communities with pathogenic properties may trigger an inadequate host inflammatory-immune response, leading to the disruption of tissue homoeostasis and development of disease. Because of their unique characteristics, MSCs are suggested as crucial regulators of tissue regeneration even under such harsh environmental conditions. The heterogeneous effects of bacteria on MSCs across studies imply the complexity underlying the interactions between stem cells and bacteria. Hence, a better understanding of stem cell behaviour at sites of inflammation appears to be a key strategy in developing new approaches for in situ tissue regeneration. Here, we review the literature on the effects of oral bacteria on cell proliferation, differentiation capacity and immunomodulation of dental-derived MSCs. PMID:26058313

  12. Alterations in radioresistance of eucaryotic cells after the transfer of genomic wildtype DNA and metallothionein genes

    International Nuclear Information System (INIS)

    The presented paper describes experiments concerning the alteration of radiosensitivity of eucaryotic cells after gene transfer. Ionizing radiation (γ- or X-ray) induces DNA single- or double strand breaks, which are religated by an unknown repair system. Repair deficient cells are highly sensitive to ionizing radiation. In the experiments described, cells from a patient with the heritable disease Ataxia telangiectasia were used as well as two X-ray sensitive CHO mutant cell lines. After gene transfer of an intact human DNA repair gene or a metallothionein gene the cells should regain radioresistance. (orig.)

  13. Unconventional Specimen Preparation Techniques Using High Resolution Low Voltage Field Emission Scanning Electron Microscopy to Study Cell Motility, Host Cell Invasion, and Internal Cell Structures in Toxoplasma gondii

    Science.gov (United States)

    Schatten, Heide; Ris, Hans

    2002-04-01

    Apicomplexan parasites employ complex and unconventional mechanisms for cell locomotion, host cell invasion, and cell division that are only poorly understood. While immunofluorescence and conventional transmission electron microscopy have been used to answer questions about the localization of some cytoskeletal proteins and cell organelles, many questions remain unanswered, partly because new methods are needed to study the complex interactions of cytoskeletal proteins and organelles that play a role in cell locomotion, host cell invasion, and cell division. The choice of fixation and preparation methods has proven critical for the analysis of cytoskeletal proteins because of the rapid turnover of actin filaments and the dense spatial organization of the cytoskeleton and its association with the complex membrane system. Here we introduce new methods to study structural aspects of cytoskeletal motility, host cell invasion, and cell division of Toxoplasma gondii, a most suitable laboratory model that is representative of apicomplexan parasites. The novel approach in our experiments is the use of high resolution low voltage field emission scanning electron microscopy (LVFESEM) combined with two new specimen preparation techniques. The first method uses LVFESEM after membrane extraction and stabilization of the cytoskeleton. This method allows viewing of actin filaments which had not been possible with any other method available so far. The second approach of imaging the parasite's ultrastructure and interactions with host cells uses semithick sections (200 nm) that are resin de-embedded (Ris and Malecki, 1993) and imaged with LVFESEM. This method allows analysis of structural detail in the parasite before and after host cell invasion and interactions with the membrane of the parasitophorous vacuole as well as parasite cell division.

  14. Proteomic profiling of SupT1 cells reveal modulation of host proteins by HIV-1 Nef variants.

    Directory of Open Access Journals (Sweden)

    Reshu Saxena

    Full Text Available Nef is an accessory viral protein that promotes HIV-1 replication, facilitating alterations in cellular pathways via multiple protein-protein interactions. The advent of proteomics has expanded the focus on better identification of novel molecular pathways regulating disease progression. In this study, nef was sequenced from randomly selected patients, however, sequence variability identified did not elicited any specific mutation that could have segregated HIV-1 patients in different stages of disease progression. To explore the difference in Nef functionality based on sequence variability we used proteomics approach. Proteomic profiling was done to compare the effect of Nef variants in host cell protein expression. 2DGE in control and Nef transfected SupT1 cells demonstrated several differentially expressed proteins. Fourteen protein spots were detected with more than 1.5 fold difference. Significant down regulation was seen in six unique protein spots in the Nef treated cells. Proteins were identified as Cyclophilin A, EIF5A-1 isoform B, Rho GDI 1 isoform a, VDAC1, OTUB1 and α-enolase isoform 1 (ENO1 through LC-MS/MS. The differential expression of the 6 proteins was analyzed by Real time PCR, Western blotting and Immunofluorescence studies with two Nef variants (RP14 and RP01 in SupT1 cells. There was contrasting difference between the effect of these Nef variants upon the expression of these six proteins. Downregulation of α-enolase (ENO1, VDAC1 and OTUB1 was more significant by Nef RP01 whereas Cyclophilin A and RhoGDI were found to be more downregulated by Nef RP14. This difference in Nef variants upon host protein expression was also studied through a site directed mutant of Nef RP01 (55AAAAAAA61 and the effect was found to be reversed. Deciphering the role of these proteins mediated by Nef variants will open a new avenue of research in understanding Nef mediated pathogenesis. Overall study determines modulation of cellular protein

  15. Rapid membrane disruption by a perforin-like protein facilitates parasite exit from host cells.

    Science.gov (United States)

    Kafsack, Björn F C; Pena, Janethe D O; Coppens, Isabelle; Ravindran, Sandeep; Boothroyd, John C; Carruthers, Vern B

    2009-01-23

    Perforin-like proteins are expressed by many bacterial and protozoan pathogens, yet little is known about their function or mode of action. Here, we describe Toxoplasma perforin-like protein 1 (TgPLP1), a secreted perforin-like protein of the intracellular protozoan pathogen Toxoplasma gondii that displays structural features necessary for pore formation. After intracellular growth, TgPLP1-deficient parasites failed to exit normally, resulting in entrapment within host cells. We show that this defect is due to an inability to rapidly permeabilize the parasitophorous vacuole membrane and host plasma membrane during exit. TgPLP1 ablation had little effect on growth in culture but resulted in a reduction greater than five orders of magnitude of acute virulence in mice. Perforin-like proteins from other intracellular pathogens may play a similar role in microbial egress and virulence. PMID:19095897

  16. Experimental approaches to investigate effector translocation into host cells in the Ustilago maydis/maize pathosystem.

    Science.gov (United States)

    Tanaka, Shigeyuki; Djamei, Armin; Presti, Libera Lo; Schipper, Kerstin; Winterberg, Sarah; Amati, Simone; Becker, Dirk; Büchner, Heike; Kumlehn, Jochen; Reissmann, Stefanie; Kahmann, Regine

    2015-01-01

    The fungus Ustilago maydis is a pathogen that establishes a biotrophic interaction with Zea mays. The interaction with the plant host is largely governed by more than 300 novel, secreted protein effectors, of which only four have been functionally characterized. Prerequisite to examine effector function is to know where effectors reside after secretion. Effectors can remain in the extracellular space, i.e. the plant apoplast (apoplastic effectors), or can cross the plant plasma membrane and exert their function inside the host cell (cytoplasmic effectors). The U. maydis effectors lack conserved motifs in their primary sequences that could allow a classification of the effectome into apoplastic/cytoplasmic effectors. This represents a significant obstacle in functional effector characterization. Here we describe our attempts to establish a system for effector classification into apoplastic and cytoplasmic members, using U. maydis for effector delivery.

  17. Early host cell targets of Yersinia pestis during primary pneumonic plague.

    Directory of Open Access Journals (Sweden)

    Roger D Pechous

    Full Text Available Inhalation of Yersinia pestis causes primary pneumonic plague, a highly lethal syndrome with mortality rates approaching 100%. Pneumonic plague progression is biphasic, with an initial pre-inflammatory phase facilitating bacterial growth in the absence of host inflammation, followed by a pro-inflammatory phase marked by extensive neutrophil influx, an inflammatory cytokine storm, and severe tissue destruction. Using a FRET-based probe to quantitate injection of effector proteins by the Y. pestis type III secretion system, we show that these bacteria target alveolar macrophages early during infection of mice, followed by a switch in host cell preference to neutrophils. We also demonstrate that neutrophil influx is unable to limit bacterial growth in the lung and is ultimately responsible for the severe inflammation during the lethal pro-inflammatory phase.

  18. Spontaneous loss and alteration of antigen receptor expression in mature CD4+ T cells

    International Nuclear Information System (INIS)

    The T-cell receptor CD3 (TCR/CD3) complex plays a central role in antigen recognition and activation of mature T cells, and therefore abnormalities in the expression of the complex should induce unresponsiveness of T cells to antigen stimulus. Using flow cytometry, we detected and enumerated variant cells with loss or alteration of surface TCR/CD3 expression among human mature CD4+ T cells. The presence of variant CD4+ T cells was demonstrated by isolating and cloning them from peripheral blood, and their abnormalities can be accounted for by alterations in TCR expression such as defects of protein expression and partial protein deletion. The variant frequency in peripheral blood increased with aging in normal donors and was highly elevated in patients with ataxia telangiectasia, an autosomal recessive inherited disease with defective DNA repair and variable T-cell immunodeficiency. These findings suggest that such alterations in TCR expression are induced by somatic mutagenesis of TCR genes and can be important factors related to age-dependent and genetic disease-associated T-cell dysfunction. (author)

  19. Bacterial porin disrupts mitochondrial membrane potential and sensitizes host cells to apoptosis.

    Directory of Open Access Journals (Sweden)

    Vera Kozjak-Pavlovic

    2009-10-01

    Full Text Available The bacterial PorB porin, an ATP-binding beta-barrel protein of pathogenic Neisseria gonorrhoeae, triggers host cell apoptosis by an unknown mechanism. PorB is targeted to and imported by host cell mitochondria, causing the breakdown of the mitochondrial membrane potential (DeltaPsi(m. Here, we show that PorB induces the condensation of the mitochondrial matrix and the loss of cristae structures, sensitizing cells to the induction of apoptosis via signaling pathways activated by BH3-only proteins. PorB is imported into mitochondria through the general translocase TOM but, unexpectedly, is not recognized by the SAM sorting machinery, usually required for the assembly of beta-barrel proteins in the mitochondrial outer membrane. PorB integrates into the mitochondrial inner membrane, leading to the breakdown of DeltaPsi(m. The PorB channel is regulated by nucleotides and an isogenic PorB mutant defective in ATP-binding failed to induce DeltaPsi(m loss and apoptosis, demonstrating that dissipation of DeltaPsi(m is a requirement for cell death caused by neisserial infection.

  20. Inhibition of gold nanoparticles (AuNPs) on pathogenic biofilm formation and invasion to host cells

    Science.gov (United States)

    Yu, Qilin; Li, Jianrong; Zhang, Yueqi; Wang, Yufan; Liu, Lu; Li, Mingchun

    2016-01-01

    Owing to the growing infectious diseases caused by eukaryotic and prokaryotic pathogens, it is urgent to develop novel antimicrobial agents against clinical pathogenic infections. Biofilm formation and invasion into the host cells are vital processes during pathogenic colonization and infection. In this study, we tested the inhibitory effect of Au nanoparticles (AuNPs) on pathogenic growth, biofilm formation and invasion. Interestingly, although the synthesized AuNPs had no significant toxicity to the tested pathogens, Candida albicans and Pseudomonas aeruginosa, the nanoparticles strongly inhibited pathogenic biofilm formation and invasion to dental pulp stem cells (DPSCs). Further investigations revealed that AuNPs abundantly bound to the pathogen cells, which likely contributed to their inhibitory effect on biofilm formation and invasion. Moreover, treatment of AuNPs led to activation of immune response-related genes in DPSCs, which may enhance the activity of host immune system against the pathogens. Zeta potential analysis and polyethylene glycol (PEG)/polyethyleneimine (PEI) coating tests further showed that the interaction between pathogen cells and AuNPs is associated with electrostatic attractions. Our findings shed novel light on the application of nanomaterials in fighting against clinical pathogens, and imply that the traditional growth inhibition test is not the only way to evaluate the drug effect during the screening of antimicrobial agents. PMID:27220400

  1. Inhibition of gold nanoparticles (AuNPs) on pathogenic biofilm formation and invasion to host cells.

    Science.gov (United States)

    Yu, Qilin; Li, Jianrong; Zhang, Yueqi; Wang, Yufan; Liu, Lu; Li, Mingchun

    2016-01-01

    Owing to the growing infectious diseases caused by eukaryotic and prokaryotic pathogens, it is urgent to develop novel antimicrobial agents against clinical pathogenic infections. Biofilm formation and invasion into the host cells are vital processes during pathogenic colonization and infection. In this study, we tested the inhibitory effect of Au nanoparticles (AuNPs) on pathogenic growth, biofilm formation and invasion. Interestingly, although the synthesized AuNPs had no significant toxicity to the tested pathogens, Candida albicans and Pseudomonas aeruginosa, the nanoparticles strongly inhibited pathogenic biofilm formation and invasion to dental pulp stem cells (DPSCs). Further investigations revealed that AuNPs abundantly bound to the pathogen cells, which likely contributed to their inhibitory effect on biofilm formation and invasion. Moreover, treatment of AuNPs led to activation of immune response-related genes in DPSCs, which may enhance the activity of host immune system against the pathogens. Zeta potential analysis and polyethylene glycol (PEG)/polyethyleneimine (PEI) coating tests further showed that the interaction between pathogen cells and AuNPs is associated with electrostatic attractions. Our findings shed novel light on the application of nanomaterials in fighting against clinical pathogens, and imply that the traditional growth inhibition test is not the only way to evaluate the drug effect during the screening of antimicrobial agents. PMID:27220400

  2. Cellular and molecular remodelling of a host cell for vertical transmission of bacterial symbionts

    Science.gov (United States)

    Luan, Jun-Bo; Shan, Hong-Wei; Isermann, Philipp; Huang, Jia-Hsin; Lammerding, Jan; Liu, Shu-Sheng; Douglas, Angela E.

    2016-01-01

    Various insects require intracellular bacteria that are restricted to specialized cells (bacteriocytes) and are transmitted vertically via the female ovary, but the transmission mechanisms are obscure. We hypothesized that, in the whitefly Bemisia tabaci, where intact bacteriocytes (and not isolated bacteria) are transferred to oocytes, the transmission mechanism would be evident as cellular and molecular differences between the nymph (pre-adult) and adult bacteriocytes. We demonstrate dramatic remodelling of bacteriocytes at the developmental transition from nymph to adulthood. This transition involves the loss of cell–cell adhesion, high division rates to constant cell size and onset of cell mobility, enabling the bacteriocytes to crawl to the ovaries. These changes are accompanied by cytoskeleton reorganization and changes in gene expression: genes functioning in cell–cell adhesion display reduced expression and genes involved in cell division, cell motility and endocytosis/exocytosis have elevated expression in adult bacteriocytes, relative to nymph bacteriocytes. This study demonstrates, for the first time, how developmentally orchestrated remodelling of gene expression and correlated changes in cell behaviour underpin the capacity of bacteriocytes to mediate the vertical transmission and persistence of the symbiotic bacteria on which the insect host depends. PMID:27358364

  3. Cell wall composition profiling of parasitic giant dodder (Cuscuta reflexa) and its hosts: a priori differences and induced changes.

    Science.gov (United States)

    Johnsen, Hanne R; Striberny, Bernd; Olsen, Stian; Vidal-Melgosa, Silvia; Fangel, Jonatan U; Willats, William G T; Rose, Jocelyn K C; Krause, Kirsten

    2015-08-01

    Host plant penetration is the gateway to survival for holoparasitic Cuscuta and requires host cell wall degradation. Compositional differences of cell walls may explain why some hosts are amenable to such degradation while others can resist infection. Antibody-based techniques for comprehensive profiling of cell wall epitopes and cell wall-modifying enzymes were applied to several susceptible hosts and a resistant host of Cuscuta reflexa and to the parasite itself. Infected tissue of Pelargonium zonale contained high concentrations of de-esterified homogalacturonans in the cell walls, particularly adjacent to the parasite's haustoria. High pectinolytic activity in haustorial extracts and high expression levels of pectate lyase genes suggest that the parasite contributes directly to wall remodeling. Mannan and xylan concentrations were low in P. zonale and in five susceptible tomato introgression lines, but high in the resistant Solanum lycopersicum cv M82, and in C. reflexa itself. Knowledge of the composition of resistant host cell walls and the parasite's own cell walls is useful in developing strategies to prevent infection by parasitic plants. PMID:25808919

  4. Probing host pathogen cross-talk by transcriptional profiling of both Mycobacterium tuberculosis and infected human dendritic cells and macrophages.

    Directory of Open Access Journals (Sweden)

    Ludovic Tailleux

    Full Text Available BACKGROUND: Transcriptional profiling using microarrays provides a unique opportunity to decipher host pathogen cross-talk on the global level. Here, for the first time, we have been able to investigate gene expression changes in both Mycobacterium tuberculosis, a major human pathogen, and its human host cells, macrophages and dendritic cells. METHODOLOGY/PRINCIPAL FINDINGS: In addition to common responses, we could identify eukaryotic and microbial transcriptional signatures that are specific to the cell type involved in the infection process. In particular M. tuberculosis shows a marked stress response when inside dendritic cells, which is in accordance with the low permissivity of these specialized phagocytes to the tubercle bacillus and to other pathogens. In contrast, the mycobacterial transcriptome inside macrophages reflects that of replicating bacteria. On the host cell side, differential responses to infection in macrophages and dendritic cells were identified in genes involved in oxidative stress, intracellular vesicle trafficking and phagosome acidification. CONCLUSIONS/SIGNIFICANCE: This study provides the proof of principle that probing the host and the microbe transcriptomes simultaneously is a valuable means to accessing unique information on host pathogen interactions. Our results also underline the extraordinary plasticity of host cell and pathogen responses to infection, and provide a solid framework to further understand the complex mechanisms involved in immunity to M. tuberculosis and in mycobacterial adaptation to different intracellular environments.

  5. Effects of bile acids on proliferation and ultrastructural alteration of pancreatic cancer cell lines

    Institute of Scientific and Technical Information of China (English)

    Zheng Wu; Yi Lüi; Bo Wang; Chang Liu; Zuo-Ren Wang,

    2003-01-01

    AIM: Pancreatic cancer in the head is frequently accompanied by jaundice and high bile acid level in serum. This study focused on the direct effects of bile acids on proliferation and ultrastructural alteration of pancreatic cancer.METHODS: Pancreatic cancer cell lines PANC-1, MIA PaCa2 and PGHAM-1 were explored in this study. The cell lines were cultured in media supplemented with certain bile acids,CA, DCA, LCA, TCDC, TDCA and GCA. Their influence on cell growth was measured with MTT assay after 72 h of incubation. Cell cycles of PANC-1 cells in 40 μM of bile acids media were analyzed by flow cytometry. Ultrastructural alteration of PANC-1 cells induced by DCA was observed using scanning and transmission electron microscope (SEM and TEM).RESULTS: At various concentrations of bile acids and incubation time, no enhanced effects of bile acids on cell proliferation were observed. Significant inhibitory effects were obtained in almost all media with bile acids. DCA and CA increased the percentage of G0+G1 phase cells, while GCA and TDCA elevated the S phase cell number. After 48 h of incubation in DCA medium, PANC-1 cells showed some structural damages such as loss of their microvilli and vacuolization of organelles in cytoplasm.CONCLUSION: Bile acids can reduce proliferation of pancreatic cancer cells due to their direct cytotoxicity. This result implies that elevation of bile acids in jaundiced serum may inhibit pancreatic cancer progression.

  6. Effects of Chemotherapy-Induced Alterations in Cell Mechanical Properties on Cancer Metastasis

    Science.gov (United States)

    Prathivadhi, Sruti; Ekpenyong, Andrew; Nichols, Michael; Taylor, Carolyn; Ning, Jianhao

    Biological cells can modulate their mechanical properties to suit their functions and in response to changes in their environment. Thus, mechanical phenotyping of cells has been employed for tracking stem cell differentiation, bacterial infection, cell death, etc. Malignant transformation of cells also involves changes in mechanical properties. However, the extent to which mechanical properties of cancer cells contribute to metastasis is not well understood. Yet, more than 90% of all cancer deaths are directly related to metastasis. Transit of cells through the microcirculation is one of the key features of metastasis. We hypothesize that cancer treatment regimens do inadvertently alter cell mechanical properties in ways that might promote cancer metastasis. We use a microfluidic microcirculation mimetic (MMM) platform which mimics the capillary constrictions of the pulmonary and peripheral microcirculation to determine if in-vivo-like mechanical stimuli can evoke different responses from cells subjected to various cancer drugs. In particular, we show that cancer cells treated with chemotherapeutic drugs such as daunorubicin, become more deformable at short timescales (0.1 s) and transit faster through the device. Our results are first steps in evaluating the pro- or anti-metastatic effects of chemotherapeutic drugs based on their induced alterations in cell mechanical properties.

  7. Tracing Conidial Fate and Measuring Host Cell Antifungal Activity Using a Reporter of Microbial Viability in the Lung

    Directory of Open Access Journals (Sweden)

    Anupam Jhingran

    2012-12-01

    Full Text Available Fluorescence can be harnessed to monitor microbial fate and to investigate functional outcomes of individual microbial cell-host cell encounters at portals of entry in native tissue environments. We illustrate this concept by introducing fluorescent Aspergillus reporter (FLARE conidia that simultaneously report phagocytic uptake and fungal viability during cellular interactions with the murine respiratory innate immune system. Our studies using FLARE conidia reveal stepwise and cell-type-specific requirements for CARD9 and Syk, transducers of C-type lectin receptor and integrin signals, in neutrophil recruitment, conidial uptake, and conidial killing in the lung. By achieving single-event resolution in defined leukocyte populations, the FLARE method enables host cell profiling on the basis of pathogen uptake and killing and may be extended to other pathogens in diverse model host organisms to query molecular, cellular, and pharmacologic mechanisms that shape host-microbe interactions.

  8. Altered cell wall properties are responsible for ammonium-reduced aluminium accumulation in rice roots.

    Science.gov (United States)

    Wang, Wei; Zhao, Xue Qiang; Chen, Rong Fu; Dong, Xiao Ying; Lan, Ping; Ma, Jian Feng; Shen, Ren Fang

    2015-07-01

    The phytotoxicity of aluminium (Al) ions can be alleviated by ammonium (NH4(+)) in rice and this effect has been attributed to the decreased Al accumulation in the roots. Here, the effects of different nitrogen forms on cell wall properties were compared in two rice cultivars differing in Al tolerance. An in vitro Al-binding assay revealed that neither NH4(+) nor NO3(-) altered the Al-binding capacity of cell walls, which were extracted from plants not previously exposed to N sources. However, cell walls extracted from NH4(+)-supplied roots displayed lower Al-binding capacity than those from NO3(-)-supplied roots when grown in non-buffered solutions. Fourier-transform infrared microspectroscopy analysis revealed that, compared with NO3(-)-supplied roots, NH4(+)-supplied roots possessed fewer Al-binding groups (-OH and COO-) and lower contents of pectin and hemicellulose. However, when grown in pH-buffered solutions, these differences in the cell wall properties were not observed. Further analysis showed that the Al-binding capacity and properties of cell walls were also altered by pHs alone. Taken together, our results indicate that the NH4(+)-reduced Al accumulation was attributed to the altered cell wall properties triggered by pH decrease due to NH4(+) uptake rather than direct competition for the cell wall binding sites between Al(3+) and NH4(+).

  9. T-cell recognition is shaped by epitope sequence conservation in the host proteome and microbiome

    DEFF Research Database (Denmark)

    Bresciani, Anne Gøther; Paul, Sinu; Schommer, Nina;

    2016-01-01

    Several mechanisms exist to avoid or suppress inflammatory T-cell immune responses that could prove harmful to the host due to targeting self-antigens or commensal microbes. We hypothesized that these mechanisms could become evident when comparing the immunogenicity of a peptide from a pathogen...... as a result of negative selection of T cells capable of recognizing such peptides. Moreover, we also found a reduced level of immune recognition for epitopes conserved in the commensal microbiome, presumably as a result of peripheral tolerance. These findings indicate that the existence (and potentially...... the polarization) of T-cell responses to a given epitope is influenced and to some extent predictable based on its similarity to self-antigens and commensal antigens....

  10. Autophagy sustains the replication of porcine reproductive and respiratory virus in host cells

    International Nuclear Information System (INIS)

    In this study, we confirmed the autophagy induced by porcine reproductive and respiratory syndrome virus (PRRSV) in permissive cells and investigated the role of autophagy in the replication of PRRSV. We first demonstrated that PRRSV infection significantly results in the increased double-membrane vesicles, the accumulation of LC3 fluorescence puncta, and the raised ratio of LC3-II/β-actin, in MARC-145 cells. Then we discovered that induction of autophagy by rapamycin significantly enhances the viral titers of PRRSV, while inhibition of autophagy by 3-MA and silencing of LC3 gene by siRNA reduces the yield of PRRSV. The results showed functional autolysosomes can be formed after PRRSV infection and the autophagosome–lysosome-fusion inhibitor decreases the virus titers. We also examined the induction of autophagy by PRRSV infection in pulmonary alveolar macrophages. These findings indicate that autophagy induced by PRRSV infection plays a role in sustaining the replication of PRRSV in host cells.

  11. Autophagy sustains the replication of porcine reproductive and respiratory virus in host cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Qinghao; Qin, Yixian; Zhou, Lei; Kou, Qiuwen; Guo, Xin; Ge, Xinna [Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine and State Key Laboratory of Agribiotechnology, China Agricultural University, Beijing (China); Yang, Hanchun, E-mail: yanghanchun1@cau.edu.cn [Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine and State Key Laboratory of Agribiotechnology, China Agricultural University, Beijing (China); Hu, Hongbo, E-mail: hongbo@cau.edu.cn [College of Food Science and Nutritional Engineering, China Agricultural University, Beijing (China)

    2012-08-01

    In this study, we confirmed the autophagy induced by porcine reproductive and respiratory syndrome virus (PRRSV) in permissive cells and investigated the role of autophagy in the replication of PRRSV. We first demonstrated that PRRSV infection significantly results in the increased double-membrane vesicles, the accumulation of LC3 fluorescence puncta, and the raised ratio of LC3-II/{beta}-actin, in MARC-145 cells. Then we discovered that induction of autophagy by rapamycin significantly enhances the viral titers of PRRSV, while inhibition of autophagy by 3-MA and silencing of LC3 gene by siRNA reduces the yield of PRRSV. The results showed functional autolysosomes can be formed after PRRSV infection and the autophagosome-lysosome-fusion inhibitor decreases the virus titers. We also examined the induction of autophagy by PRRSV infection in pulmonary alveolar macrophages. These findings indicate that autophagy induced by PRRSV infection plays a role in sustaining the replication of PRRSV in host cells.

  12. A Review: competence, compromise, and concomitance-reaction of the host cell to toxoplasma gondii infection and development

    OpenAIRE

    Peng, H. J.; Chen, X. G.; Lindsay, D S

    2011-01-01

    Toxoplasma gondii is an important zoonotic parasite with a worldwide distribution. It infects about one-third of the world's population, causing serious illness in immunosuppressed individuals, fetuses, and infants. Toxoplasma gondii biology within the host cell includes several important phases: (I) active invasion and establishment of a nonfusogenic parasitophorous vacuole in the host cell, (2) extensive modification of the parasitophorous vacuolar membrane for nutrient acquisition, (3) int...

  13. The Influence of Virus Infection on the Extracellular pH of the Host Cell Detected on Cell Membrane.

    Science.gov (United States)

    Liu, Hengjun; Maruyama, Hisataka; Masuda, Taisuke; Honda, Ayae; Arai, Fumihito

    2016-01-01

    Influenza virus infection can result in changes in the cellular ion levels at 2-3 h post-infection. More H(+) is produced by glycolysis, and the viral M2 proton channel also plays a role in the capture and release of H(+) during both viral entry and egress. Then the cells might regulate the intracellular pH by increasing the export of H(+) from the intracellular compartment. Increased H(+) export could lead indirectly to increased extracellular acidity. To detect changes in extracellular pH of both virus-infected and uninfected cells, pH sensors were synthesized using polystyrene beads (ϕ1 μm) containing Rhodamine B and Fluorescein isothiocyanate (FITC). The fluorescence intensity of FITC can respond to both pH and temperature. So Rhodamine B was also introduced in the sensor for temperature compensation. Then the pH can be measured after temperature compensation. The sensor was adhered to cell membrane for extracellular pH measurement. The results showed that the multiplication of influenza virus in host cell decreased extracellular pH of the host cell by 0.5-0.6 in 4 h after the virus bound to the cell membrane, compared to that in uninfected cells. Immunostaining revealed the presence of viral PB1 protein in the nucleus of virus-bound cells that exhibited extracellular pH changes, but no PB1 protein are detected in virus-unbound cells where the extracellular pH remained constant. PMID:27582727

  14. Radiation-induced alterations in murine lymphocyte homing patterns. II. Recovery and function of memory cells in LBN rats

    International Nuclear Information System (INIS)

    Suspensions of lymph node cells from dinitrophenylated bovine gamma globulin (DNP-BGG)-immune LBN F1 hybrid rats (Lewis X Brown Norway) were prepared, irradiated, and injected intravenously into unirradiated syngeneic intermediate hosts and irradiated syngeneic adoptive controls. After allowance of 24 hr for homing to occur, the intermediate hosts were killed and cell preparations from the lymph nodes and spleen were injected intravenously into separate irradiated LBN final host groups. All control and experimental groups were challenged (DNP-BGG saline iv) 24 hr after the injection of the lymphoid cells. Rats were bled on Days 7, 11, and 14 after challenge and the antigen-binding capacity (ABC) of the serum was determined. After correction for the fraction of the total cell population transferred from the intermediate host, the peak ABC of the final hosts was related to the number of memory cells present. It was thus possible to determine the relative distribution of the memory cell population to the spleen and lymph nodes of the intermediate hosts. In the intermediate control animals, irradiated memory cells provided a secondary antibody response which was delayed but not suppressed when compared to unirradiated cells. In intermediate hosts, the homing of lymph node memory cells to the spleen and lymph nodes was significantly reduced by an exposure to 200 R of x radiation

  15. Vibrio cholerae Porin OmpU Induces Caspase-independent Programmed Cell Death upon Translocation to the Host Cell Mitochondria.

    Science.gov (United States)

    Gupta, Shelly; Prasad, G V R Krishna; Mukhopadhaya, Arunika

    2015-12-25

    Porins, a major class of outer membrane proteins in Gram-negative bacteria, primarily act as transport channels. OmpU is one of the major porins of human pathogen, Vibrio cholerae. In the present study, we show that V. cholerae OmpU has the ability to induce target cell death. Although OmpU-mediated cell death shows some characteristics of apoptosis, such as flipping of phosphatidylserine in the membrane as well as cell size shrinkage and increased cell granularity, it does not show the caspase-3 activation and DNA laddering pattern typical of apoptotic cells. Increased release of lactate dehydrogenase in OmpU-treated cells indicates that the OmpU-mediated cell death also has characteristics of necrosis. Further, we show that the mechanism of OmpU-mediated cell death involves major mitochondrial changes in the target cells. We observe that OmpU treatment leads to the disruption of mitochondrial membrane potential, resulting in the release of cytochrome c and apoptosis-inducing factor (AIF). AIF translocates to the host cell nucleus, implying that it has a crucial role in OmpU-mediated cell death. Finally, we observe that OmpU translocates to the target cell mitochondria, where it directly initiates mitochondrial changes leading to mitochondrial membrane permeability transition and AIF release. Partial blocking of AIF release by cyclosporine A in OmpU-treated cells further suggests that OmpU may be inducing the opening of the mitochondrial permeability transition pore. All of these results lead us to the conclusion that OmpU induces cell death in target cells in a programmed manner in which mitochondria play a central role.

  16. Tumor-altered dendritic cell function: implications for anti-tumor immunity

    Directory of Open Access Journals (Sweden)

    Kristian Michael Hargadon

    2013-07-01

    Full Text Available Dendritic cells are key regulators of both innate and adaptive immunity, and the array of immunoregulatory functions exhibited by these cells is dictated by their differentiation, maturation, and activation status. Although a major role for these cells in the induction of immunity to pathogens has long been appreciated, data accumulated over the last several years has demonstrated that DC are also critical regulators of anti-tumor immune responses. However, despite the potential for stimulation of robust anti-tumor immunity by DC, tumor-altered DC function has been observed in many cancer patients and tumor-bearing animals and is often associated with tumor immune escape. Such dysfunction has significant implications for both the induction of natural anti-tumor immune responses as well as the efficacy of immunotherapeutic strategies that target endogenous DC in situ or that employ exogenous DC as part of anti-cancer immunization maneuvers. In this review, the major types of tumor-altered DC function will be described, with emphasis on recent insights into the mechanistic bases for the inhibition of DC differentiation from hematopoietic precursors, the altered programming of DC precursors to differentiate into myeloid-derived suppressor cells or tumor-associated macrophages, the suppression of DC maturation and activation, and the induction of immunoregulatory DC by tumors, tumor-derived factors, and tumor-associated cells within the milieu of the tumor microenvironment. The impact of these tumor-altered cells on the quality of the overall anti-tumor immune response will also be discussed. Finally, this review will also highlight questions concerning tumor-altered DC function that remain unanswered, and it will address factors that have limited advances in the study of this phenomenon in order to focus future research efforts in the field on identifying strategies for interfering with tumor-associated DC dysfunction and improving DC-mediated anti

  17. Cell structure and cytokinesis alterations in multidrug-resistant Leishmania (Leishmania) amazonensis.

    Science.gov (United States)

    Borges, V M; Lopes, U G; De Souza, W; Vannier-Santos, M A

    2005-01-01

    Multidrug-resistant Leishmania (Leishmania) amazonensis may be obtained by in vitro selection with vinblastine. In order to determine whether this phenotype is linked to structural alterations, we analyzed the cell architecture by electron microscopy. The vinblastine resistant CL2 clone of L. (L.) amazonensis, but not wild-type parasites, showed a cytokinesis dysfunction. The CL2 promastigotes had multiple nuclei, kinetoplasts and flagella, suggesting that vinblastine resistance may be associated with truncated cell division. The subpellicular microtubule plasma membrane connection was also affected. Wild-type parasites treated with vinblastine displayed similar alterations, presenting lobulated and multinucleated cells. Taken together, these data indicate that antimicrotubule drug-selected parasites may show evidence of the mutation of cytoskeleton proteins, impairing normal cell function. PMID:15592939

  18. Quantitative analysis of the supernatant from host and transfected CHO cells using iTRAQ 8-plex technique.

    Science.gov (United States)

    Zhu, Guijie; Sun, Liangliang; Albanetti, Thomas; Linkous, Travis; Larkin, Christopher; Schoner, Ronald; McGivney, James B; Dovichi, Norman J

    2016-10-01

    We employed UPLC-MS/MS with iTRAQ 8-plex labeling to quantitatively analyze the supernatant produced by two Chinese hamster ovary (CHO) cell lines (CHO K1SV and CHO CAT-S). In each case, the supernatant from the host and three transfected clones were analyzed at days 5, 7, and 10 of culture. A total of eight iTRAQ 8-plex experiments were performed. For each cell line, the overlap of supernatant protein identifications between transfected clones is over 60%. Over 70% of the supernatant proteins in the CHO K1SV host cell line are present in the CHO CAT-S cell line. For the CHO K1SV cell line, the overlap in supernatant protein identifications between the host cell line and the transfected clones is >59%. For the CHO CAT-S cell line, the overlap between supernatant protein identifications for the transfected clone and host cell is >45%. These differences in the supernatant protein identifications between transfected clones in each cell line and between the two host cell lines are not significant. We used cluster analysis to characterize the change in supernatant protein expression as a function of cell culture time. Roughly 1.3 or clones at each time point. Greater than 65% of the common proteins in the CHO K1SV cell line supernatant and over 54% in the CHO CAT-S cell line supernatant show no significant expression difference between host and the three transfected clones. Data are available via ProteomeXchange with identifier PXD003462. Biotechnol. Bioeng. 2016;113: 2140-2148. © 2016 Wiley Periodicals, Inc. PMID:27070921

  19. Lethal graft-versus-host disease: modification with allogeneic cultured donor cells

    International Nuclear Information System (INIS)

    The use of the bone marrow culture technique was studied as a means to prepare donor marrow for bone marrow transplantation to avoid lethal graft-versus-host disease (GVHD). Preliminary experiments demonstrated the rapid loss of theta-positive cells in such cultures, so that theta-positive cells were not detected after 6 days. Initial experiments in C3H/HeJ (H-2k, Hbbd) recipients prepared with 900 rad demonstrated improved survival when 3-day cultured C57BL/6 (H-2b, Hbbs) donor cells were used in place of hind limb marrow for transplantation. However, hemoglobin typing of recipient animals revealed only short-term donor engraftment, with competitive repopulation of recipient marrow occurring. Subsequent experiments were done in 1,200-rad prepared recipients, with long-term donor engraftment demonstrated. The majority of 1,200-rad prepared animals receiving cultured allogeneic cells died of GVHD, but animals receiving 28-day cultured cells had an improved 90-day survival and a delay in GVHD development over animals receiving hind limb marrow or marrow from shorter times in culture. In addition, animals receiving anti-theta-treated, 3-day nonadherent cells had an improved survival (44%) over animals receiving anti-theta-treated hind limb marrow (20%). These experiments demonstrate modest benefit for the use of cultured cells in bone marrow transplantation across major H-2 histocompatibility complex differences

  20. Human gastric mucins differently regulate Helicobacter pylori proliferation, gene expression and interactions with host cells.

    Directory of Open Access Journals (Sweden)

    Emma C Skoog

    Full Text Available Helicobacter pylori colonizes the mucus niche of the gastric mucosa and is a risk factor for gastritis, ulcers and cancer. The main components of the mucus layer are heavily glycosylated mucins, to which H. pylori can adhere. Mucin glycosylation differs between individuals and changes during disease. Here we have examined the H. pylori response to purified mucins from a range of tumor and normal human gastric tissue samples. Our results demonstrate that mucins from different individuals differ in how they modulate both proliferation and gene expression of H. pylori. The mucin effect on proliferation varied significantly between samples, and ranged from stimulatory to inhibitory, depending on the type of mucins and the ability of the mucins to bind to H. pylori. Tumor-derived mucins and mucins from the surface mucosa had potential to stimulate proliferation, while gland-derived mucins tended to inhibit proliferation and mucins from healthy uninfected individuals showed little effect. Artificial glycoconjugates containing H. pylori ligands also modulated H. pylori proliferation, albeit to a lesser degree than human mucins. Expression of genes important for the pathogenicity of H. pylori (babA, sabA, cagA, flaA and ureA appeared co-regulated in response to mucins. The addition of mucins to co-cultures of H. pylori and gastric epithelial cells protected the viability of the cells and modulated the cytokine production in a manner that differed between individuals, was partially dependent of adhesion of H. pylori to the gastric cells, but also revealed that other mucin factors in addition to adhesion are important for H. pylori-induced host signaling. The combined data reveal host-specific effects on proliferation, gene expression and virulence of H. pylori due to the gastric mucin environment, demonstrating a dynamic interplay between the bacterium and its host.

  1. Adjustment of host cells for accommodation of symbiotic bacteria: vacuole defunctionalization, HOPS suppression, and TIP1g retargeting in Medicago

    NARCIS (Netherlands)

    Gavrin, A.Y.; Kaiser, B.N.; Geiger, D.; Tyerman, S.D.; Wen, Z.; Bisseling, T.; Fedorova, E.E.

    2014-01-01

    In legume–rhizobia symbioses, the bacteria in infected cells are enclosed in a plant membrane, forming organelle-like compartments called symbiosomes. Symbiosomes remain as individual units and avoid fusion with lytic vacuoles of host cells. We observed changes in the vacuole volume of infected cell

  2. Cutting edge: IL-17-secreting innate lymphoid cells are essential for host defense against fungal infection.

    Science.gov (United States)

    Gladiator, André; Wangler, Nicolette; Trautwein-Weidner, Kerstin; LeibundGut-Landmann, Salomé

    2013-01-15

    IL-17-mediated immunity has emerged as a crucial host defense mechanism against fungal infections. Although Th cells are generally thought to act as the major source of IL-17 in response to Candida albicans, we show that fungal control is mediated by IL-17-secreting innate lymphoid cells (ILCs) and not by Th17 cells. By using a mouse model of oropharyngeal candidiasis we found that IL-17A and IL-17F, which are both crucial for pathogen clearance, are produced promptly upon infection in an IL-23-dependent manner, and that ILCs in the oral mucosa are the main source for these cytokines. Ab-mediated depletion of ILCs in RAG1-deficient mice or ILC deficiency in retinoic acid-related orphan receptor c(-/-) mice resulted in a complete failure to control the infection. Taken together, our data uncover the cellular basis for the IL-23/IL-17 axis, which acts right at the onset of infection when it is most needed for fungal control and host protection.

  3. The Host Cell Receptors for Measles Virus and Their Interaction with the Viral Hemagglutinin (H Protein

    Directory of Open Access Journals (Sweden)

    Liang-Tzung Lin

    2016-09-01

    Full Text Available The hemagglutinin (H protein of measles virus (MeV interacts with a cellular receptor which constitutes the initial stage of infection. Binding of H to this host cell receptor subsequently triggers the F protein to activate fusion between virus and host plasma membranes. The search for MeV receptors began with vaccine/laboratory virus strains and evolved to more relevant receptors used by wild-type MeV. Vaccine or laboratory strains of measles virus have been adapted to grow in common cell lines such as Vero and HeLa cells, and were found to use membrane cofactor protein (CD46 as a receptor. CD46 is a regulator that normally prevents cells from complement-mediated self-destruction, and is found on the surface of all human cells, with the exception of erythrocytes. Mutations in the H protein, which occur during adaptation and allow the virus to use CD46 as a receptor, have been identified. Wild-type isolates of measles virus cannot use the CD46 receptor. However, both vaccine/laboratory and wild-type strains can use an immune cell receptor called signaling lymphocyte activation molecule family member 1 (SLAMF1; also called CD150 and a recently discovered epithelial receptor known as Nectin-4. SLAMF1 is found on activated B, T, dendritic, and monocyte cells, and is the initial target for infections by measles virus. Nectin-4 is an adherens junction protein found at the basal surfaces of many polarized epithelial cells, including those of the airways. It is also over-expressed on the apical and basal surfaces of many adenocarcinomas, and is a cancer marker for metastasis and tumor survival. Nectin-4 is a secondary exit receptor which allows measles virus to replicate and amplify in the airways, where the virus is expelled from the body in aerosol droplets. The amino acid residues of H protein that are involved in binding to each of the receptors have been identified through X-ray crystallography and site-specific mutagenesis. Recombinant measles

  4. Over-expression and localization of a host protein on the membrane of Cryptosporidium parvum infected epithelial cells.

    Science.gov (United States)

    Yang, Yi-Lin; Serrano, Myrna G; Sheoran, Abhineet S; Manque, Patricio A; Buck, Gregory A; Widmer, Giovanni

    2009-11-01

    The genus Cryptosporidium includes several species of intestinal protozoan parasites which multiply in intestinal epithelial cells. The impact of this infection on the transcriptome of cultured host cells was investigated using DNA microarray hybridizations. The expression of 14 genes found to be consistently up- or down-regulated in infected cell monolayers was validated with RT PCR. Using immunofluorescence we examined the expression of Protease Activated Receptor-2, which is encoded by one of the up-regulated genes. In infected cells this receptor localized to the host cell membrane which covers the intracellular trophozoites and meronts. This observation indicates that the composition of the host cell membrane is affected by the developing trophozoite, a phenomenon which has not been described previously.

  5. Heat shock gene expression and cytoskeletal alterations in mouse neuroblastoma cells

    NARCIS (Netherlands)

    Bergen en Henegouwen, P.M.P. van; Linnemans, W.A.M.

    1987-01-01

    The cytoskeleton of neuroblastoma cells, clone Neuro 2A, is altered by two stress conditions: heat shock and arsenite treatment. Microtubules are reorganized, intermediate filaments are aggregated around the nucleus, and the number of stress fibers is reduced. Since both stress modalities induce sim

  6. Detection of mitochondrial deoxyribonucleic acid alterations in urine from urothelial cell carcinoma patients.

    NARCIS (Netherlands)

    Dasgupta, S.; Shao, C.; Keane, T.E.; Duberow, D.P.; Mathies, R.A.; Fisher, P.B.; Kiemeney, L.A.L.M.; Sidransky, D.

    2012-01-01

    Our study aims at understanding the timing and nature of mitochondrial deoxyribonucleic acid (mtDNA) alterations in urothelial cell carcinoma (UCC) and their detection in urine sediments. The entire 16.5 kb mitochondrial genome was sequenced in matched normal lymphocytes, tumor and urine sediments f

  7. Immune Mechanisms of Mesenchymal Stem Cell Therapy for Acute Graft versus Host Disease

    OpenAIRE

    Tobin, Laura M.

    2012-01-01

    The aim of this work was to investigate the role of Mesenchymal stem cells (MSC) in the modulation of immune responses, focusing on suppression and induction of immune tolerance. To date, MSC therapy has proved beneficial for the treatment of inflammatory and autoimmune diseases, such as acute Graft versus Host Disease (aGvHD) and Crohn’s disease. However, the exact mechanisms of therapeutic benefit remain unclear. The key goals of this study were to identify the role of MSC derived soluble a...

  8. Sucrose synthase affects carbon partitioning to increase cellulose production and altered cell wall ultrastructure

    OpenAIRE

    Coleman, Heather D.; Yan, Jimmy; Mansfield, Shawn D

    2009-01-01

    Overexpression of the Gossypium hirsutum sucrose synthase (SuSy) gene under the control of 2 promoters was examined in hybrid poplar (Populus alba × grandidentata). Analysis of RNA transcript abundance, enzyme activity, cell wall composition, and soluble carbohydrates revealed significant changes in the transgenic lines. All lines showed significantly increased SuSy enzyme activity in developing xylem. This activity manifested in altered secondary cell wall cellulose content per dry weight in...

  9. The pluralization of the international: Resistance and alter-standardization in regenerative stem cell medicine

    OpenAIRE

    Rosemann, Achim; Chaisinthop, Nattaka

    2016-01-01

    The article explores the formation of an international politics of resistance and ‘alter-standardization’ in regenerative stem cell medicine. The absence of internationally harmonized regulatory frameworks in the clinical stem cell field and the presence of lucrative business opportunities have resulted in the formation of transnational networks adopting alternative research standards and practices. These oppose, as a universal global standard, strict evidence-based medicine clinical research...

  10. δ-Catenin promotes prostate cancer cell growth and progression by altering cell cycle and survival gene profiles

    Directory of Open Access Journals (Sweden)

    Chen Yan-Hua

    2009-03-01

    Full Text Available Abstract Background δ-Catenin is a unique member of β-catenin/armadillo domain superfamily proteins and its primary expression is restricted to the brain. However, δ-catenin is upregulated in human prostatic adenocarcinomas, although the effects of δ-catenin overexpression in prostate cancer are unclear. We hypothesized that δ-catenin plays a direct role in prostate cancer progression by altering gene profiles of cell cycle regulation and cell survival. Results We employed gene transfection and small interfering RNA to demonstrate that increased δ-catenin expression promoted, whereas its knockdown suppressed prostate cancer cell viability. δ-Catenin promoted prostate cancer cell colony formation in soft agar as well as tumor xenograft growth in nude mice. Deletion of either the amino-terminal or carboxyl-terminal sequences outside the armadillo domains abolished the tumor promoting effects of δ-catenin. Quantitative RT2 Profiler™ PCR Arrays demonstrated gene alterations involved in cell cycle and survival regulation. δ-Catenin overexpression upregulated cyclin D1 and cdc34, increased phosphorylated histone-H3, and promoted the entry of mitosis. In addition, δ-catenin overexpression resulted in increased expression of cell survival genes Bcl-2 and survivin while reducing the cell cycle inhibitor p21Cip1. Conclusion Taken together, our studies suggest that at least one consequence of an increased expression of δ-catenin in human prostate cancer is the alteration of cell cycle and survival gene profiles, thereby promoting tumor progression.

  11. Herpes Simplex Virus 1 Infection Alters the mRNA Translation Processing in L-02 Cells

    Institute of Scientific and Technical Information of China (English)

    Min HONG; Yan-chun CHE; Gui-zhen TANG; Wei CUN; Xue-mei ZHANG; Long-ding LIU; Qi-han LI

    2008-01-01

    HSV-1 infection-mediated regulation of mRNA translation in host cells is a systematic and complicated process. Investigation of the details of this mechanism will facilitate understanding of biological variations in the viral replication process and host cells. In this study, a comparative proteomics technology platform was applied by two-dimension electrophoresis of HSV-1 infected normal human L-02 cell and control cell lysates. The observed protein spots were analyzed qualitatively and quantitatively by the PDQuest software package. A number of the different observed protein spots closely associated with cellular protein synthesis were identified by matrix-assisted laser-desorption ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The expression levels of the RPLP1 protein, which is required for mRNA translation, and KHSRP protein, which is involved in rapid decay of mRNA, were up-regulated, whereas the expression level of RNP H2, which is involved in positive regulation on the mRNA splicing process, was down-regulated. All of these results suggest that HSV-1 infection can influence cellular protein synthesis via modulation of cellular regulatory proteins involved in RNA splicing, translation and decay, resulting in optimisation of viral protein synthesis when cellular protein synthesis is shut off. Although there is need for further investigations regarding the detailed mechanisms of cellular protein control, our studies provide new insight into the targeting of varied virus signaling pathways involved in host cellular protein synthesis.

  12. Tc17 cells mediate vaccine immunity against lethal fungal pneumonia in immune deficient hosts lacking CD4+ T cells.

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    Som Gowda Nanjappa

    Full Text Available Vaccines may help reduce the growing incidence of fungal infections in immune-suppressed patients. We have found that, even in the absence of CD4(+ T-cell help, vaccine-induced CD8(+ T cells persist and confer resistance against Blastomyces dermatitidis and Histoplasma capsulatum. Type 1 cytokines contribute to that resistance, but they also are dispensable. Although the role of T helper 17 cells in immunity to fungi is debated, IL-17 producing CD8(+ T cells (Tc17 cells have not been investigated. Here, we show that Tc17 cells are indispensable in antifungal vaccine immunity in hosts lacking CD4(+ T cells. Tc17 cells are induced upon vaccination, recruited to the lung on pulmonary infection, and act non-redundantly in mediating protection in a manner that requires neutrophils. Tc17 cells did not influence type I immunity, nor did the lack of IL-12 signaling augment Tc17 cells, indicating a distinct lineage and function. IL-6 was required for Tc17 differentiation and immunity, but IL-1R1 and Dectin-1 signaling was unexpectedly dispensable. Tc17 cells expressed surface CXCR3 and CCR6, but only the latter was essential in recruitment to the lung. Although IL-17 producing T cells are believed to be short-lived, effector Tc17 cells expressed low levels of KLRG1 and high levels of the transcription factor TCF-1, predicting their long-term survival and stem-cell like behavior. Our work has implications for designing vaccines against fungal infections in immune suppressed patients.

  13. Tc17 cells mediate vaccine immunity against lethal fungal pneumonia in immune deficient hosts lacking CD4+ T cells.

    Science.gov (United States)

    Nanjappa, Som Gowda; Heninger, Erika; Wüthrich, Marcel; Gasper, David Joseph; Klein, Bruce S

    2012-01-01

    Vaccines may help reduce the growing incidence of fungal infections in immune-suppressed patients. We have found that, even in the absence of CD4(+) T-cell help, vaccine-induced CD8(+) T cells persist and confer resistance against Blastomyces dermatitidis and Histoplasma capsulatum. Type 1 cytokines contribute to that resistance, but they also are dispensable. Although the role of T helper 17 cells in immunity to fungi is debated, IL-17 producing CD8(+) T cells (Tc17 cells) have not been investigated. Here, we show that Tc17 cells are indispensable in antifungal vaccine immunity in hosts lacking CD4(+) T cells. Tc17 cells are induced upon vaccination, recruited to the lung on pulmonary infection, and act non-redundantly in mediating protection in a manner that requires neutrophils. Tc17 cells did not influence type I immunity, nor did the lack of IL-12 signaling augment Tc17 cells, indicating a distinct lineage and function. IL-6 was required for Tc17 differentiation and immunity, but IL-1R1 and Dectin-1 signaling was unexpectedly dispensable. Tc17 cells expressed surface CXCR3 and CCR6, but only the latter was essential in recruitment to the lung. Although IL-17 producing T cells are believed to be short-lived, effector Tc17 cells expressed low levels of KLRG1 and high levels of the transcription factor TCF-1, predicting their long-term survival and stem-cell like behavior. Our work has implications for designing vaccines against fungal infections in immune suppressed patients.

  14. Extracellular superoxide dismutase protects Histoplasma yeast cells from host-derived oxidative stress.

    Directory of Open Access Journals (Sweden)

    Brian H Youseff

    Full Text Available In order to establish infections within the mammalian host, pathogens must protect themselves against toxic reactive oxygen species produced by phagocytes of the immune system. The fungal pathogen Histoplasma capsulatum infects both neutrophils and macrophages but the mechanisms enabling Histoplasma yeasts to survive in these phagocytes have not been fully elucidated. We show that Histoplasma yeasts produce a superoxide dismutase (Sod3 and direct it to the extracellular environment via N-terminal and C-terminal signals which promote its secretion and association with the yeast cell surface. This localization permits Sod3 to protect yeasts specifically from exogenous superoxide whereas amelioration of endogenous reactive oxygen depends on intracellular dismutases such as Sod1. While infection of resting macrophages by Histoplasma does not stimulate the phagocyte oxidative burst, interaction with polymorphonuclear leukocytes (PMNs and cytokine-activated macrophages triggers production of reactive oxygen species (ROS. Histoplasma yeasts producing Sod3 survive co-incubation with these phagocytes but yeasts lacking Sod3 are rapidly eliminated through oxidative killing similar to the effect of phagocytes on Candida albicans yeasts. The protection provided by Sod3 against host-derived ROS extends in vivo. Without Sod3, Histoplasma yeasts are attenuated in their ability to establish respiratory infections and are rapidly cleared with the onset of adaptive immunity. The virulence of Sod3-deficient yeasts is restored in murine hosts unable to produce superoxide due to loss of the NADPH-oxidase function. These results demonstrate that phagocyte-produced ROS contributes to the immune response to Histoplasma and that Sod3 facilitates Histoplasma pathogenesis by detoxifying host-derived reactive oxygen thereby enabling Histoplasma survival.

  15. Yersinia pseudotuberculosis Spatially Controls Activation and Misregulation of Host Cell Rac1.

    Directory of Open Access Journals (Sweden)

    2005-10-01

    Full Text Available Yersinia pseudotuberculosis binds host cells and modulates the mammalian Rac1 guanosine triphosphatase (GTPase at two levels. Activation of Rac1 results from integrin receptor engagement, while misregulation is promoted by translocation of YopE and YopT proteins into target cells. Little is known regarding how these various factors interplay to control Rac1 dynamics. To investigate these competing processes, the localization of Rac1 activation was imaged microscopically using fluorescence resonance energy transfer. In the absence of translocated effectors, bacteria induced activation of the GTPase at the site of bacterial binding. In contrast, the entire cellular pool of Rac1 was inactivated shortly after translocation of YopE RhoGAP. Inactivation required membrane localization of Rac1. The translocated protease YopT had very different effects on Rac1. This protein, which removes the membrane localization site of Rac1, did not inactivate Rac1, but promoted entry of cleaved activated Rac1 molecules into the host cell nucleus, allowing Rac1 to localize with nuclear guanosine nucleotide exchange factors. As was true for YopE, membrane-associated Rac1 was the target for YopT, indicating that the two translocated effectors may compete for the same pool of target protein. Consistent with the observation that YopE inactivation requires membrane localization of Rac1, the presence of YopT in the cell interfered with the action of the YopE RhoGAP. As a result, interaction of target cells with a strain that produces both YopT and YopE resulted in two spatially distinct pools of Rac1: an inactive cytoplasmic pool and an activated nuclear pool. These studies demonstrate that competition between bacterial virulence factors for access to host substrates is controlled by the spatial arrangement of a target protein. In turn, the combined effects of translocated bacterial proteins are to generate pools of a single signaling molecule with distinct localization and

  16. Adipose Tissue-Derived Mesenchymal Stem Cells as a New Host Cell in Latent Leishmaniasis

    OpenAIRE

    Allahverdiyev, Adil M; Bagirova, Melahat; Elcicek, Serhat; Koc, Rabia Cakir; Baydar, Serap Yesilkir; Findikli, Necati; Oztel, Olga N.

    2011-01-01

    Some protozoan infections such as Toxoplasma, Cryptosporidium, and Plasmodium can be transmitted through stem cell transplantations. To our knowledge, so far, there is no study about transmission of Leishmania parasites in stem cell transplantation and interactions between parasites and stem cells in vitro. Therefore, the aim of this study was to investigate the interaction between different species of Leishmania parasites and adipose tissue-derived mesenchymal stem cells (ADMSCs). ADMSCs hav...

  17. Alterations in Cell-Extracellular Matrix Interactions during Progression of Cancers

    Directory of Open Access Journals (Sweden)

    Rajeswari Jinka

    2012-01-01

    Full Text Available Cancer progression is a multistep process during which normal cells exhibit molecular changes that culminate into the highly malignant and metastatic phenotype, observed in cancerous tissues. The initiation of cell transformation is generally associated with genetic alterations in normal cells that lead to the loss of intercellular- and/or extracellular-matrix- (ECM- mediated cell adhesion. Transformed cells undergo rapid multiplication and generate more modifications in adhesion and motility-related molecules which allow them to escape from the original site and acquire invasive characteristics. Integrins, which are multifunctional adhesion receptors, and are present, on normal as well as transformed cells, assist the cells undergoing tumor progression in creating the appropriate environment for their survival, growth, and invasion. In this paper, we have briefly discussed the role of ECM proteins and integrins during cancer progression and described some unique conditions where adhesion-related changes could induce genetic mutations in anchorage-independent tumor model systems.

  18. Prenatal immune activation alters hippocampal place cell firing characteristics in adult animals.

    Science.gov (United States)

    Wolff, Amy R; Bilkey, David K

    2015-08-01

    Prenatal maternal immune activation (MIA) is a risk factor for several developmental neuropsychiatric disorders, including autism, bipolar disorder and schizophrenia. Adults with these disorders display alterations in memory function that may result from changes in the structure and function of the hippocampus. In the present study we use an animal model to investigate the effect that a transient prenatal maternal immune activation episode has on the spatially-modulated firing activity of hippocampal neurons in adult animals. MIA was induced in pregnant rat dams with a single injection of the synthetic cytokine inducer polyinosinic:polycytidylic acid (poly I:C) on gestational day 15. Control dams were given a saline equivalent. Firing activity and local field potentials (LFPs) were recorded from the CA1 region of the adult male offspring of these dams as they moved freely in an open arena. Most neurons displayed characteristic spatially-modulated 'place cell' firing activity and while there was no between-group difference in mean firing rate between groups, place cells had smaller place fields in MIA-exposed animals when compared to control-group cells. Cells recorded in MIA-group animals also displayed an altered firing-phase synchrony relationship to simultaneously recorded LFPs. When the floor of the arena was rotated, the place fields of MIA-group cells were more likely to shift in the same direction as the floor rotation, suggesting that local cues may have been more salient for these animals. In contrast, place fields in control group cells were more likely to shift firing position to novel spatial locations suggesting an altered response to contextual cues. These findings show that a single MIA intervention is sufficient to change several important characteristics of hippocampal place cell activity in adult offspring. These changes could contribute to the memory dysfunction that is associated with MIA, by altering the encoding of spatial context and by

  19. Alteration of cadherin isoform expression and inhibition of gap junctions in stomach carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    To explore cell malignant phenotype correlated changes of cell surface adhesion molecules and cell-cell communication in carcinogenesis, human stomach transformed and cancer cell lines were investigated. Expressions of E-cadherin, N-cadherin, ?-catenin, ?-catenin as well as gap junction (GJ) protein Cx32 were studied by utilization of immunoblotting, immunocytochemical and fluorescent dye transfer methods. Mammalian normal stomach mucosal cells expressed E-cadherin but not N-cadherin. E-cadherin immunofluorescence was detected at cell membranous adherens junctions (AJ) where colocalization with immunofluorescent staining of inner surface adhesion plaque proteins ?- and ?-catenins was observed. The existence of E-cadherin/ catenin (?-, ?-) protein complexes as AJ was suggested. In transformed and stomach cancer cells E-cadherin was inhibited, instead, N-cadherin was expressed and localized at membranous AJ where co-staining with ?- and ?-catenin fluorescence was observed. Formation of N-cadherin/catenin (?-, ?-) protein complex at AJs of transformed and cancer cells was suggested. The above observations were further supported by immunoblotting results. Normal stomach muscosal and transformed cells expressed Cx32 at membranous GJ and were competent of gap junction communication (GJIC). In stomach cancer cells, Cx32 was inhibited and GJIC was defective. The results suggested that changes of signal pathways mediated by both cell adhesion and cell communication systems are associated intracellular events of stomach carcinogenesis. The alteration of cadherin isoform from E- to N-cadherin in transformed and stomach cancer cells is the first report.

  20. More Novel Hantaviruses and Diversifying Reservoir Hosts — Time for Development of Reservoir-Derived Cell Culture Models?

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    Isabella Eckerle

    2014-02-01

    Full Text Available Due to novel, improved and high-throughput detection methods, there is a plethora of newly identified viruses within the genus Hantavirus. Furthermore, reservoir host species are increasingly recognized besides representatives of the order Rodentia, now including members of the mammalian orders Soricomorpha/Eulipotyphla and Chiroptera. Despite the great interest created by emerging zoonotic viruses, there is still a gross lack of in vitro models, which reflect the exclusive host adaptation of most zoonotic viruses. The usually narrow host range and genetic diversity of hantaviruses make them an exciting candidate for studying virus-host interactions on a cellular level. To do so, well-characterized reservoir cell lines covering a wide range of bat, insectivore and rodent species are essential. Most currently available cell culture models display a heterologous virus-host relationship and are therefore only of limited value. Here, we review the recently established approaches to generate reservoir-derived cell culture models for the in vitro study of virus-host interactions. These successfully used model systems almost exclusively originate from bats and bat-borne viruses other than hantaviruses. Therefore we propose a parallel approach for research on rodent- and insectivore-borne hantaviruses, taking the generation of novel rodent and insectivore cell lines from wildlife species into account. These cell lines would be also valuable for studies on further rodent-borne viruses, such as orthopox- and arenaviruses.

  1. Transforming growth factor-β2 induces morphological alteration of human corneal endothelial cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Jing; Wang; Ting-Jun; Fan; Xiu-Xia; Yang; Shi-Min; Chang

    2014-01-01

    AIM:To investigate the morphological altering effect of transforming growth factor-β2(TGF-β2) on untransfected human corneal endothelial cells(HCECs)in vitro.METHODS:After untransfected HCECs were treated with TGF-β2 at different concentrations, the morphology,cytoskeleton distribution, and type IV collagen expression of the cells were examined with inverted contrast light microscopy, fluorescence microscopy,immunofluorescence or Western Blot.RESULTS:TGF-β2 at the concentration of 3-15 μg/L had obviously alterative effects on HCECs morphology in dose and time-dependent manner, and 9 μg/L was the peak concentration. TGF-β2(9 μg/L) altered HCE cell morphology after treatment for 36 h, increased the mean optical density(P <0.01) and the length of F-actin,reduced the mean optical density(P <0.01) of the collagen type IV in extracellular matrix(ECM) and induced the rearrangement of F-actin, microtubule in cytoplasm and collagen type IV in ECM after treatment for 72 h.·CONCLUTION: TGF-β2 has obviously alterative effect on the morphology of HCECs from polygonal phenotype to enlarged spindle-shaped phenotype, in dose and time-dependence manner by inducing more, elongation and alignment of F-actin, rearrangement of microtubule and larger spread area of collagen type IV.

  2. Modulation of Cell Sialoglycophenotype: A Stylish Mechanism Adopted by Trypanosoma cruzi to Ensure Its Persistence in the Infected Host

    Science.gov (United States)

    Freire-de-Lima, Leonardo; da Fonseca, Leonardo M.; da Silva, Vanessa A.; da Costa, Kelli M.; Morrot, Alexandre; Freire-de-Lima, Célio G.; Previato, Jose O.; Mendonça-Previato, Lucia

    2016-01-01

    Trypanosoma cruzi, the etiological agent of Chagas disease exhibits multiple mechanisms to guarantee its establishment and persistence in the infected host. It has been well demonstrated that T. cruzi is not able to synthesize sialic acids (Sia). To acquire the monosaccharide, the parasite makes use of a multifunctional enzyme called trans-sialidase (Tc-TS). Since this enzyme has no analogous in the vertebrate host, it has been used as a target in drug therapy development. Tc-TS preferentially catalyzes the transfer of Sia from the host glycoconjugates to the terminal β-galactopyranosyl residues of mucin-like molecules present on the parasite’s cell surface. Alternatively, the enzyme can sialylate/re-sialylate glycoconjugates expressed on the surface of host cells. Since its discovery, several studies have shown that T. cruzi employs the Tc-TS activity to modulate the host cell sialoglycophenotype, thus favoring its perpetuation in the infected vertebrate. In this review, we summarize the dynamic of host/parasite sialoglycophenotype modulation, highlighting its role in the subversion of host immune response in order to promote the establishment of persistent chronic infection. PMID:27242722

  3. Purification of infectious human herpesvirus 6A virions and association of host cell proteins

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    Garoff Henrik

    2007-10-01

    Full Text Available Abstract Background Viruses that are incorporating host cell proteins might trigger autoimmune diseases. It is therefore of interest to identify possible host proteins associated with viruses, especially for enveloped viruses that have been suggested to play a role in autoimmune diseases, like human herpesvirus 6A (HHV-6A in multiple sclerosis (MS. Results We have established a method for rapid and morphology preserving purification of HHV-6A virions, which in combination with parallel analyses with background control material released from mock-infected cells facilitates qualitative and quantitative investigations of the protein content of HHV-6A virions. In our iodixanol gradient purified preparation, we detected high levels of viral DNA by real-time PCR and viral proteins by metabolic labelling, silver staining and western blots. In contrast, the background level of cellular contamination was low in the purified samples as demonstrated by the silver staining and metabolic labelling analyses. Western blot analyses showed that the cellular complement protein CD46, the receptor for HHV-6A, is associated with the purified and infectious virions. Also, the cellular proteins clathrin, ezrin and Tsg101 are associated with intact HHV-6A virions. Conclusion Cellular proteins are associated with HHV-6A virions. The relevance of the association in disease and especially in autoimmunity will be further investigated.

  4. Lactobacilli Interfere with Streptococcus pyogenes Hemolytic Activity and Adherence to Host Epithelial Cells.

    Science.gov (United States)

    Saroj, Sunil D; Maudsdotter, Lisa; Tavares, Raquel; Jonsson, Ann-Beth

    2016-01-01

    Streptococcus pyogenes [Group A streptococcus (GAS)], a frequent colonizer of the respiratory tract mucosal surface, causes a variety of human diseases, ranging from pharyngitis to the life-threatening streptococcal toxic shock-like syndrome. Lactobacilli have been demonstrated to colonize the respiratory tract. In this study, we investigated the interference of lactobacilli with the virulence phenotypes of GAS. The Lactobacillus strains L. rhamnosus Kx151A1 and L. reuteri PTA-5289, but not L. salivarius LMG9477, inhibited the hemolytic activity of S. pyogenes S165. The inhibition of hemolytic activity was attributed to a decrease in the production of streptolysin S (SLS). Conditioned medium (CM) from the growth of L. rhamnosus Kx151A1 and L. reuteri PTA-5289 was sufficient to down-regulate the expression of the sag operon, encoding SLS. The Lactobacillus strains L. rhamnosus Kx151A1, L. reuteri PTA-5289, and L. salivarius LMG9477 inhibited the initial adherence of GAS to host epithelial cells. Intriguingly, competition with a combination of Lactobacillus species reduced GAS adherence to host cells most efficiently. The data suggest that an effector molecule released from certain Lactobacillus strains attenuates the production of SLS at the transcriptional level and that combinations of Lactobacillus strains may protect the pharyngeal mucosa more efficiently from the initial colonization of GAS. The effector molecules released from Lactobacillus strains affecting the virulence phenotypes of pathogens hold potential in the development of a new generation of therapeutics.

  5. Lactobacilli Interfere with Streptococcus pyogenes Hemolytic Activity and Adherence to Host Epithelial Cells.

    Science.gov (United States)

    Saroj, Sunil D; Maudsdotter, Lisa; Tavares, Raquel; Jonsson, Ann-Beth

    2016-01-01

    Streptococcus pyogenes [Group A streptococcus (GAS)], a frequent colonizer of the respiratory tract mucosal surface, causes a variety of human diseases, ranging from pharyngitis to the life-threatening streptococcal toxic shock-like syndrome. Lactobacilli have been demonstrated to colonize the respiratory tract. In this study, we investigated the interference of lactobacilli with the virulence phenotypes of GAS. The Lactobacillus strains L. rhamnosus Kx151A1 and L. reuteri PTA-5289, but not L. salivarius LMG9477, inhibited the hemolytic activity of S. pyogenes S165. The inhibition of hemolytic activity was attributed to a decrease in the production of streptolysin S (SLS). Conditioned medium (CM) from the growth of L. rhamnosus Kx151A1 and L. reuteri PTA-5289 was sufficient to down-regulate the expression of the sag operon, encoding SLS. The Lactobacillus strains L. rhamnosus Kx151A1, L. reuteri PTA-5289, and L. salivarius LMG9477 inhibited the initial adherence of GAS to host epithelial cells. Intriguingly, competition with a combination of Lactobacillus species reduced GAS adherence to host cells most efficiently. The data suggest that an effector molecule released from certain Lactobacillus strains attenuates the production of SLS at the transcriptional level and that combinations of Lactobacillus strains may protect the pharyngeal mucosa more efficiently from the initial colonization of GAS. The effector molecules released from Lactobacillus strains affecting the virulence phenotypes of pathogens hold potential in the development of a new generation of therapeutics. PMID:27524981

  6. Epigenetic alteration of imprinted genes during neural differentiation of germline-derived pluripotent stem cells.

    Science.gov (United States)

    Lee, Hye Jeong; Choi, Na Young; Lee, Seung-Won; Ko, Kisung; Hwang, Tae Sook; Han, Dong Wook; Lim, Jisun; Schöler, Hans R; Ko, Kinarm

    2016-03-01

    Spermatogonial stem cells (SSCs), which are unipotent stem cells in the testes that give rise to sperm, can be converted into germline-derived pluripotent stem (gPS) by self-induction. The androgenetic imprinting pattern of SSCs is maintained even after their reprogramming into gPS cells. In this study, we used an in vitro neural differentiation model to investigate whether the imprinting patterns are maintained or altered during differentiation. The androgenetic patterns of H19, Snrpn, and Mest were maintained even after differentiation of gPS cells into NSCs (gPS-NSCs), whereas the fully unmethylated status of Ndn in SSCs was altered to somatic patterns in gPS cells and gPS-NSCs. Thus, our study demonstrates epigenetic alteration of genomic imprinting during the induction of pluripotency in SSCs and neural differentiation, suggesting that gPS-NSCs can be a useful model to study the roles of imprinted genes in brain development and human neurodevelopmental disorders.

  7. Subcellular distribution of swine vesicular disease virus proteins and alterations induced in infected cells: A comparative study with foot-and-mouth disease virus and vesicular stomatitis virus

    International Nuclear Information System (INIS)

    The intracellular distribution of swine vesicular disease virus (SVDV) proteins and the induced reorganization of endomembranes in IBRS-2 cells were analyzed. Fluorescence to new SVDV capsids appeared first upon infection, concentrated in perinuclear circular structures and colocalized to dsRNA. As in foot-and-mouth disease virus (FMDV)-infected cells, a vesicular pattern was predominantly found in later stages of SVDV capsid morphogenesis that colocalized with those of non-structural proteins 2C, 2BC and 3A. These results suggest that assembly of capsid proteins is associated to the replication complex. Confocal microscopy showed a decreased fluorescence to ER markers (calreticulin and protein disulfide isomerase), and disorganization of cis-Golgi gp74 and trans-Golgi caveolin-1 markers in SVDV- and FMDV-, but not in vesicular stomatitis virus (VSV)-infected cells. Electron microscopy of SVDV-infected cells at an early stage of infection revealed fragmented ER cisternae with expanded lumen and accumulation of large Golgi vesicles, suggesting alterations of vesicle traffic through Golgi compartments. At this early stage, FMDV induced different patterns of ER fragmentation and Golgi alterations. At later stages of SVDV cytopathology, cells showed a completely vacuolated cytoplasm containing vesicles of different sizes. Cell treatment with brefeldin A, which disrupts the Golgi complex, reduced SVDV (∼ 5 log) and VSV (∼ 4 log) titers, but did not affect FMDV growth. Thus, three viruses, which share target tissues and clinical signs in natural hosts, induce different intracellular effects in cultured cells

  8. The effect of Bulgarian propolis against Trypanosoma cruzi and during its interaction with host cells

    Directory of Open Access Journals (Sweden)

    Andréia Pires Dantas

    2006-03-01

    Full Text Available Propolis has shown activity against pathogenic microorganisms that cause diseases in humans and animals. The ethanol (Et-Blg and acetone (Ket-Blg extracts from a Bulgarian propolis, with known chemical compositions, presented similar activity against tissue culture-derived amastigotes. The treatment of Trypanosoma cruzi-infected skeletal muscle cells with Et-Blg led to a decrease of infection and of the intracellular proliferation of amastigotes, while damage to the host cell was observed only at concentration 12.5 times higher than those affecting the parasite. Ultrastructural analysis of the effect of both extracts in epimastigotes revealed that the main targets were the mitochondrion and reservosomes. Et-Blg also affected the mitochondrion-kinetoplast complex in trypomastigotes, offering a potential target for chemotherapeutic agents.

  9. Role of skin immune cells on the host susceptibility to mosquito-borne viruses.

    Science.gov (United States)

    Briant, Laurence; Desprès, Philippe; Choumet, Valérie; Missé, Dorothée

    2014-09-01

    Due to climate change and the propagation of competent arthropods worldwide, arboviruses have become pathogens of major medical importance. Early transmission to vertebrates is initiated by skin puncture and deposition of virus together with arthropod saliva in the epidermis and dermis. Saliva components have the capacity to modulate skin cell responses by enhancing and/or counteracting initial replication and establishment of systemic viral infection. Here, we review the nature of the cells targeted by arboviruses at the skin level and discuss the type of cellular responses elicited by these pathogens in light of the immunomodulatory properties of arthropod vector-derived salivary factors injected at the inoculation site. Understanding cutaneous arbovirus-host interactions may provide new clues for the design of future therapeutics. PMID:25043586

  10. Altered goblet cell differentiation and surface mucus properties in Hirschsprung disease.

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    Jay R Thiagarajah

    Full Text Available Hirschsprung disease-associated enterocolitis (HAEC leads to significant mortality and morbidity, but its pathogenesis remains unknown. Changes in the colonic epithelium related to goblet cells and the luminal mucus layer have been postulated to play a key role. Here we show that the colonic epithelium of both aganglionic and ganglionic segments are altered in patients and in mice with Hirschsprung disease (HSCR. Structurally, goblet cells were altered with increased goblet cell number and reduced intracellular mucins in the distal colon of biopsies from patients with HSCR. Endothelin receptor B (Ednrb mutant mice showed increased goblet cell number and size and increased cell proliferation compared to wild-type mice in aganglionic segments, and reduced goblet cell size and number in ganglionic segments. Functionally, compared to littermates, Ednrb-/- mice showed increased transepithelial resistance, reduced stool water content and similar chloride secretion in the distal colon. Transcript levels of goblet cell differentiation factors SPDEF and Math1 were increased in the distal colon of Ednrb-/- mice. Both distal colon from Ednrb mice and biopsies from HSCR patients showed reduced Muc4 expression as compared to controls, but similar expression of Muc2. Particle tracking studies showed that mucus from Ednrb-/- mice provided a more significant barrier to diffusion of 200 nm nanoparticles as compared to wild-type mice. These results suggest that aganglionosis is associated with increased goblet cell proliferation and differentiation and subsequent altered surface mucus properties, prior to the development of inflammation in the distal colon epithelium. Restoration of normal goblet cell function and mucus layer properties in the colonic epithelium may represent a therapeutic strategy for prevention of HAEC.

  11. Cell Cycle Control and Adhesion Molecule Expression in Cells of the Immune System are Sensitive to Altered Gravity

    Science.gov (United States)

    Ullrich, O.; Paulsen, K.; Thiel, C.; Herrmann, K.; Sang, C.; Han, G.; Hemmersbach, R.; von der Wiesche, M.; Kroll, H.; Zhuang, F.; Grote, K. H.; Cogoli, A.; Zipp, F.; Engelmann, F.

    2008-06-01

    Life on earth developed in the presence and under the constant influence of gravity. Thus, it is a fundamental biological question, whether gravity is required for cellular functions and signal transduction in mammalian cells. Since the first Spacelab-Mission 20 years ago, it is known that activation and function of T lymphocytes is severely suppressed in microgravity, but the underlying molecular mechanisms are not elucidated. Experiments have been performed using ground-based facilities such as fast-rotating clinostat and hyper-g-centrifuges, and real microgravity provided by parabolic flights. We found that 1.) cells of the immune system responded cell type specifically to altered gravity, 2.) microgravity induced a multitude of initial alterations in signal transduction, whereas 3.) hypergravity of 1.8g did not induce any changes of the pathways tested, and that 4.) most of the initially altered pathways in microgravity adapted to "normal" levels within 15min. However, some pathways remained altered and could explain cell cycle arrest of T lymphocytes as observed in several long-term space experiments.

  12. Coronavirus spike protein inhibits host cell translation by interaction with eIF3f.

    Directory of Open Access Journals (Sweden)

    Han Xiao

    Full Text Available In response to viral infection, the expression of numerous host genes, including predominantly a number of proinflammatory cytokines and chemokines, is usually up-regulated at both transcriptional and translational levels. It was noted that in cells infected with coronavirus, transcription and translation of some of these genes were differentially induced. Drastic induction of their expression at the transcriptional level was observed in cells infected with coronavirus. However, induction of the same genes at the translational level was usually found to be minimal to moderate. To investigate the underlying mechanisms, yeast two-hybrid screen was carried out using SARS-CoV proteins as baits, revealing that a subunit of the eukaryotic initiation factor 3 (eIF3, eIF3f, may interact with the N-terminal region of the SARS-CoV spike (S protein. This interaction was subsequently confirmed by co-immunoprecipitation and immunofluorescent staining. Meanwhile, parallel experiments confirmed that eIF3f could also interact with the S protein of another coronavirus, the avian coronavirus infectious bronchitis virus (IBV. These interactions led to the inhibition of translation of a reporter gene in both in vitro expression system and intact cells. Interestingly, IBV-infected cells stably expressing a Flag-tagged eIF3f showed much higher translation of IL-6 and IL-8, suggesting that the interaction between coronavirus S protein and eIF3f plays a functional role in controlling the expression of host genes, especially genes that are induced during coronavirus infection cycles. This study reveals a novel mechanism exploited by coronavirus to regulate viral pathogenesis.

  13. Membrane cholesterol regulates lysosome-plasma membrane fusion events and modulates Trypanosoma cruzi invasion of host cells.

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    Bárbara Hissa

    Full Text Available BACKGROUND: Trypomastigotes of Trypanosoma cruzi are able to invade several types of non-phagocytic cells through a lysosomal dependent mechanism. It has been shown that, during invasion, parasites trigger host cell lysosome exocytosis, which initially occurs at the parasite-host contact site. Acid sphingomyelinase released from lysosomes then induces endocytosis and parasite internalization. Lysosomes continue to fuse with the newly formed parasitophorous vacuole until the parasite is completely enclosed by lysosomal membrane, a process indispensable for a stable infection. Previous work has shown that host membrane cholesterol is also important for the T. cruzi invasion process in both professional (macrophages and non-professional (epithelial phagocytic cells. However, the mechanism by which cholesterol-enriched microdomains participate in this process has remained unclear. METHODOLOGY/PRINCIPAL FINDING: In the present work we show that cardiomyocytes treated with MβCD, a drug able to sequester cholesterol from cell membranes, leads to a 50% reduction in invasion by T. cruzi trypomastigotes, as well as a decrease in the number of recently internalized parasites co-localizing with lysosomal markers. Cholesterol depletion from host membranes was accompanied by a decrease in the labeling of host membrane lipid rafts, as well as excessive lysosome exocytic events during the earlier stages of treatment. Precocious lysosomal exocytosis in MβCD treated cells led to a change in lysosomal distribution, with a reduction in the number of these organelles at the cell periphery, and probably compromises the intracellular pool of lysosomes necessary for T. cruzi invasion. CONCLUSION/SIGNIFICANCE: Based on these results, we propose that cholesterol depletion leads to unregulated exocytic events, reducing lysosome availability at the cell cortex and consequently compromise T. cruzi entry into host cells. The results also suggest that two different pools of

  14. Altered insulin receptor signalling and β-cell cycle dynamics in type 2 diabetes mellitus.

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    Franco Folli

    Full Text Available Insulin resistance, reduced β-cell mass, and hyperglucagonemia are consistent features in type 2 diabetes mellitus (T2DM. We used pancreas and islets from humans with T2DM to examine the regulation of insulin signaling and cell-cycle control of islet cells. We observed reduced β-cell mass and increased α-cell mass in the Type 2 diabetic pancreas. Confocal microscopy, real-time PCR and western blotting analyses revealed increased expression of PCNA and down-regulation of p27-Kip1 and altered expression of insulin receptors, insulin receptor substrate-2 and phosphorylated BAD. To investigate the mechanisms underlying these findings, we examined a mouse model of insulin resistance in β-cells--which also exhibits reduced β-cell mass, the β-cell-specific insulin receptor knockout (βIRKO. Freshly isolated islets and β-cell lines derived from βIRKO mice exhibited poor cell-cycle progression, nuclear restriction of FoxO1 and reduced expression of cell-cycle proteins favoring growth arrest. Re-expression of insulin receptors in βIRKO β-cells reversed the defects and promoted cell cycle progression and proliferation implying a role for insulin-signaling in β-cell growth. These data provide evidence that human β- and α-cells can enter the cell-cycle, but proliferation of β-cells in T2DM fails due to G1-to-S phase arrest secondary to defective insulin signaling. Activation of insulin signaling, FoxO1 and proteins in β-cell-cycle progression are attractive therapeutic targets to enhance β-cell regeneration in the treatment of T2DM.

  15. Toxicity of drinking water disinfection byproducts: cell cycle alterations induced by the monohaloacetonitriles.

    Science.gov (United States)

    Komaki, Yukako; Mariñas, Benito J; Plewa, Michael J

    2014-10-01

    Haloacetonitriles (HANs) are a chemical class of drinking water disinfection byproducts (DBPs) that form from reactions between disinfectants and nitrogen-containing precursors, the latter more prevalent in water sources impacted by algae bloom and municipal wastewater effluent discharge. HANs, previously demonstrated to be genotoxic, were investigated for their effects on the mammalian cell cycle. Treating Chinese hamster ovary (CHO) cells with monoHANs followed by the release from the chemical treatment resulted in the accumulation of abnormally high DNA content in cells over time (hyperploid). The potency for the cell cycle alteration followed the order: iodoacetonitrile (IAN) > bromoacetonitrile (BAN) ≫ chloroacetonitrile (CAN). Exposure to 6 μM IAN, 12 μM BAN and 900 μM CAN after 26 h post-treatment incubation resulted in DNA repair; however, subsequent cell cycle alteration effects were observed. Cell proliferation of HAN-treated cells was suppressed for as long as 43 to 52 h. Enlarged cell size was observed after 52 h post-treatment incubation without the induction of cytotoxicity. The HAN-mediated cell cycle alteration was mitosis- and proliferation-dependent, which suggests that HAN treatment induced mitosis override, and that HAN-treated cells proceeded into S phase and directly into the next cell cycle. Cells with multiples genomes would result in aneuploidy (state of abnormal chromosome number and DNA content) at the next mitosis since extra centrosomes could compromise the assembly of bipolar spindles. There is accumulating evidence of a transient tetraploid state proceeding to aneuploidy in cancer progression. Biological self-defense systems to ensure genomic stability and to eliminate tetraploid cells exist in eukaryotic cells. A key tumor suppressor gene, p53, is oftentimes mutated in various types of human cancer. It is possible that HAN disruption of the normal cell cycle and the generation of aberrant cells with an abnormal number of

  16. Alteration of natural killer(NK) cells in atomic bomb survivors of hiroshima

    International Nuclear Information System (INIS)

    This paper reports on the alteration of natural killer(NK) cells and their responsiveness to IL-2 observed in 125 atomic-bomb survivors. It is found no difference in the number and activity of NK cells among different dose groups with the same age ATB. But there was of difference in NK activity in different age ATB groups with same dose, especially in the g roups 25 years, the old with doses of 0.01-1 Gy (P < 0.05). This result suggests that there is an obvious late effect of ionizing radiation on activity of NK cells in children

  17. Analysis of Borrelia burgdorferi surface proteins as determinants in establishing host cell interactions

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    Virginia L Schmit

    2011-07-01

    Full Text Available Borrelia burgdorferi infection causes Lyme borreliosis in humans, a condition which can involve a systemic spread of the organism to colonize various tissues and organs. If the infection is left untreated by antimicrobials, it can lead to manifestations including, arthritis, carditis, and/or neurological problems. Identification and characterization of B. burgdorferi outer membrane proteins that facilitate cellular attachment and invasion to establish infection continue to be investigated. In this study, we sought to further define putative cell binding properties of surface-exposed B. burgdorferi proteins by observing whether cellular adherence could be blocked by antibodies. B. burgdorferi mixed separately with monoclonal antibodies against outer surface protein (Osp A, OspC, decorin-binding protein (Dbp A, BBA64, and RevA antigens were incubated with human umbilical vein endothelial cells (HUVEC and human neuroglial cells (H4. B. burgdorferi treated with anti-OspA, -DbpA, and –BBA64 monoclonal antibodies showed a significant decrease in cellular association compared to controls, whereas B. burgdorferi treated with anti-OspC and anti-RevA showed no reduction in cellular attachment. Additionally, temporal transcriptional analyses revealed upregulated expression of bba64, ospA, and dbpA during coincubation with cells. Together, the data provide evidence that OspA, DbpA, and BBA64 function in host cell adherence and infection mechanisms.

  18. Host cell Golgi anti-apoptotic protein (GAAP) and growth of Chlamydia pneumoniae.

    Science.gov (United States)

    Markkula, Eveliina; Hulkkonen, Jaakko; Penttilä, Tuula; Puolakkainen, Mirja

    2013-01-01

    Chlamydia pneumoniae protein CPn0809 is a type three secretion system substrate, the exact function of which in infection pathogenesis has remained unknown. In this study, we identified by yeast two-hybrid screening a potential host cell interaction partner of CPn0809, Golgi anti-apoptotic protein (GAAP), a conserved protein found in eukaryotic cells. GAAP gene is expressed at relatively constant levels and its expression remained stable also after C. pneumoniae infection. The interaction between GAAP and C. pneumoniae was suggested by transfection studies. GAAP knock-down by siRNA in infected A549 cells resulted in an increased number of C. pneumoniae genomes and growth of the bacteria as judged by quantitative PCR and inclusion counts, respectively. Silencing of GAAP did not make the A549 cells more susceptible to apoptosis per se, and infection with C. pneumoniae prevented staurosporin-induced apoptosis also in transfected cultures. Taken together, the proposed interaction between C. pneumoniae and GAAP modulates bacterial growth in A549 cells. PMID:23000903

  19. Which Way In? The RalF Arf-GEF Orchestrates Rickettsia Host Cell Invasion.

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    Kristen E Rennoll-Bankert

    2015-08-01

    Full Text Available Bacterial Sec7-domain-containing proteins (RalF are known only from species of Legionella and Rickettsia, which have facultative and obligate intracellular lifestyles, respectively. L. pneumophila RalF, a type IV secretion system (T4SS effector, is a guanine nucleotide exchange factor (GEF of ADP-ribosylation factors (Arfs, activating and recruiting host Arf1 to the Legionella-containing vacuole. In contrast, previous in vitro studies showed R. prowazekii (Typhus Group RalF is a functional Arf-GEF that localizes to the host plasma membrane and interacts with the actin cytoskeleton via a unique C-terminal domain. As RalF is differentially encoded across Rickettsia species (e.g., pseudogenized in all Spotted Fever Group species, it may function in lineage-specific biology and pathogenicity. Herein, we demonstrate RalF of R. typhi (Typhus Group interacts with the Rickettsia T4SS coupling protein (RvhD4 via its proximal C-terminal sequence. RalF is expressed early during infection, with its inactivation via antibody blocking significantly reducing R. typhi host cell invasion. For R. typhi and R. felis (Transitional Group, RalF ectopic expression revealed subcellular localization with the host plasma membrane and actin cytoskeleton. Remarkably, R. bellii (Ancestral Group RalF showed perinuclear localization reminiscent of ectopically expressed Legionella RalF, for which it shares several structural features. For R. typhi, RalF co-localization with Arf6 and PI(4,5P2 at entry foci on the host plasma membrane was determined to be critical for invasion. Thus, we propose recruitment of PI(4,5P2 at entry foci, mediated by RalF activation of Arf6, initiates actin remodeling and ultimately facilitates bacterial invasion. Collectively, our characterization of RalF as an invasin suggests that, despite carrying a similar Arf-GEF unknown from other bacteria, different intracellular lifestyles across Rickettsia and Legionella species have driven divergent roles

  20. Which Way In? The RalF Arf-GEF Orchestrates Rickettsia Host Cell Invasion.

    Science.gov (United States)

    Rennoll-Bankert, Kristen E; Rahman, M Sayeedur; Gillespie, Joseph J; Guillotte, Mark L; Kaur, Simran J; Lehman, Stephanie S; Beier-Sexton, Magda; Azad, Abdu F

    2015-08-01

    Bacterial Sec7-domain-containing proteins (RalF) are known only from species of Legionella and Rickettsia, which have facultative and obligate intracellular lifestyles, respectively. L. pneumophila RalF, a type IV secretion system (T4SS) effector, is a guanine nucleotide exchange factor (GEF) of ADP-ribosylation factors (Arfs), activating and recruiting host Arf1 to the Legionella-containing vacuole. In contrast, previous in vitro studies showed R. prowazekii (Typhus Group) RalF is a functional Arf-GEF that localizes to the host plasma membrane and interacts with the actin cytoskeleton via a unique C-terminal domain. As RalF is differentially encoded across Rickettsia species (e.g., pseudogenized in all Spotted Fever Group species), it may function in lineage-specific biology and pathogenicity. Herein, we demonstrate RalF of R. typhi (Typhus Group) interacts with the Rickettsia T4SS coupling protein (RvhD4) via its proximal C-terminal sequence. RalF is expressed early during infection, with its inactivation via antibody blocking significantly reducing R. typhi host cell invasion. For R. typhi and R. felis (Transitional Group), RalF ectopic expression revealed subcellular localization with the host plasma membrane and actin cytoskeleton. Remarkably, R. bellii (Ancestral Group) RalF showed perinuclear localization reminiscent of ectopically expressed Legionella RalF, for which it shares several structural features. For R. typhi, RalF co-localization with Arf6 and PI(4,5)P2 at entry foci on the host plasma membrane was determined to be critical for invasion. Thus, we propose recruitment of PI(4,5)P2 at entry foci, mediated by RalF activation of Arf6, initiates actin remodeling and ultimately facilitates bacterial invasion. Collectively, our characterization of RalF as an invasin suggests that, despite carrying a similar Arf-GEF unknown from other bacteria, different intracellular lifestyles across Rickettsia and Legionella species have driven divergent roles for Ral

  1. Stromal-epithelial interactions in aging and cancer: Senescent fibroblasts alter epithelial cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Parrinello, Simona; Coppe, Jean-Philippe; Krtolica, Ana; Campisi, Judith

    2004-07-14

    Cellular senescence suppresses cancer by arresting cells at risk for malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation. Moreover, using cultured mouse or human fibroblasts and non-malignant breast epithelial cells, we show that senescent fibroblasts disrupt epithelial alveolar morphogenesis, functional differentiation, and branching morphogenesis. Further, we identify MMP-3 as the major factor responsible for the effects of senescent fibroblasts on branching morphogenesis. Our findings support the idea that senescent cells contribute to age-related pathology, including cancer, and describe a new property of senescent fibroblasts--the ability to alter epithelial differentiation--that might also explain the loss of tissue function and organization that is a hallmark of aging.

  2. Helicobacter pylori Disrupts Host Cell Membranes, Initiating a Repair Response and Cell Proliferation

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    Hsueh-Fen Juan

    2012-08-01

    Full Text Available Helicobacter pylori (H. pylori, the human stomach pathogen, lives on the inner surface of the stomach and causes chronic gastritis, peptic ulcer, and gastric cancer. Plasma membrane repair response is a matter of life and death for human cells against physical and biological damage. We here test the hypothesis that H. pylori also causes plasma membrane disruption injury, and that not only a membrane repair response but also a cell proliferation response are thereby activated. Vacuolating cytotoxin A (VacA and cytotoxin-associated gene A (CagA have been considered to be major H. pylori virulence factors. Gastric cancer cells were infected with H. pylori wild type (vacA+/cagA+, single mutant (ΔvacA or ΔcagA or double mutant (ΔvacA/ΔcagA strains and plasma membrane disruption events and consequent activation of membrane repair components monitored. H. pylori disrupts the host cell plasma membrane, allowing localized dye and extracellular Ca2+ influx. Ca2+-triggered members of the annexin family, A1 and A4, translocate, in response to injury, to the plasma membrane, and cell surface expression of an exocytotic maker of repair, LAMP-2, increases. Additional forms of plasma membrane disruption, unrelated to H. pylori exposure, also promote host cell proliferation. We propose that H. pylori activation of a plasma membrane repair is pro-proliferative. This study might therefore provide new insight into potential mechanisms of H. pylori-induced gastric carcinogenesis.

  3. Mast Cell Stabilizers as Host Modulatory Drugs to Prevent and Control Periodontal Disease

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    Dhoom Singh Mehta

    2011-01-01

    Full Text Available Introduction: Mast cells are among the first cells to get in-volved in periodontal inflammation. Their numbers have been shown to be in-creased in cases of gingivitis and periodontal disease. The hypothesis: Since mast cell stabilizers like sodium cromogly-cate (SCG and nedocromil sodium (NS have been used in the prophylaxis of bronchial asthma without any significant adverse effects and also the fact that drugs like SCG show significant anti-inflammatory activities, it would be logical to use mast cell stabilizers as host modulating drugs for the treatment and prevention of peri-odontal disease. Evaluation of the hypothesis: Safety and efficacy of both SCG and NS are well documented. So, it will be systemically safe to use in humans. However, oral administration SCG or delivery of the drug by means local irrigation will not be very useful because SCG may not be secreted in the gingival crevicular fluid (GCF(as in the case of oral administraion or the drug may get washed out from periodontal pocket due to the constant flow of GCF(as in the case of irrigation. A local or targeted drug delivery of mast cell stabilizers can be used in patients with periodontal disease. Role of mast cells in periodontal disease has been dealt in-depth in many studies and articles. However, limited amount of research has been done on using mast cell stabilizers in the prevention and control of periodontal diseases. More studies are needed to study the efficacy and effective-ness of mast cell stabilizers as an adjunct to phase I therapy in the control of periodontal disease.

  4. Prolonged Mitosis of Neural Progenitors Alters Cell Fate in the Developing Brain.

    Science.gov (United States)

    Pilaz, Louis-Jan; McMahon, John J; Miller, Emily E; Lennox, Ashley L; Suzuki, Aussie; Salmon, Edward; Silver, Debra L

    2016-01-01

    Embryonic neocortical development depends on balanced production of progenitors and neurons. Genetic mutations disrupting progenitor mitosis frequently impair neurogenesis; however, the link between altered mitosis and cell fate remains poorly understood. Here we demonstrate that prolonged mitosis of radial glial progenitors directly alters neuronal fate specification and progeny viability. Live imaging of progenitors from a neurogenesis mutant, Magoh(+/-), reveals that mitotic delay significantly correlates with preferential production of neurons instead of progenitors, as well as apoptotic progeny. Independently, two pharmacological approaches reveal a causal relationship between mitotic delay and progeny fate. As mitotic duration increases, progenitors produce substantially more apoptotic progeny or neurons. We show that apoptosis, but not differentiation, is p53 dependent, demonstrating that these are distinct outcomes of mitotic delay. Together our findings reveal that prolonged mitosis is sufficient to alter fates of radial glia progeny and define a new paradigm to understand how mitosis perturbations underlie brain size disorders such as microcephaly.

  5. Altering adsorbed proteins or cellular gene expression in bone-metastatic cancer cells affects PTHrP and Gli2 without altering cell growth

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    Jonathan M. Page

    2015-09-01

    Full Text Available The contents of this data in brief are related to the article titled “Matrix Rigidity Regulates the Transition of Tumor Cells to a Bone-Destructive Phenotype through Integrin β3 and TGF-β Receptor Type II”. In this DIB we will present our supplemental data investigating Integrin expression, attachment of cells to various adhesion molecules, and changes in gene expression in multiple cancer cell lines. Since the interactions of Integrins with adsorbed matrix proteins are thought to affect the ability of cancer cells to interact with their underlying substrates, we examined the expression of Integrin β1, β3, and β5 in response to matrix rigidity. We found that only Iβ3 increased with increasing substrate modulus. While it was shown that fibronectin greatly affects the expression of tumor-produced factors associated with bone destruction (parathyroid hormone-related protein, PTHrP, and Gli2, poly-l-lysine, vitronectin and type I collagen were also analyzed as potential matrix proteins. Each of the proteins was independently adsorbed on both rigid and compliant polyurethane films which were subsequently used to culture cancer cells. Poly-l-lysine, vitronectin and type I collagen all had negligible effects on PTHrP or Gli2 expression, but fibronectin was shown to have a dose dependent effect. Finally, altering the expression of Iβ3 demonstrated that it is required for tumor cells to respond to the rigidity of the matrix, but does not affect other cell growth or viability. Together these data support the data presented in our manuscript to show that the rigidity of bone drives Integrinβ3/TGF-β crosstalk, leading to increased expression of Gli2 and PTHrP.

  6. Low Doses of Cisplatin Induce Gene Alterations, Cell Cycle Arrest, and Apoptosis in Human Promyelocytic Leukemia Cells.

    Science.gov (United States)

    Velma, Venkatramreddy; Dasari, Shaloam R; Tchounwou, Paul B

    2016-01-01

    Cisplatin is a known antitumor drug, but its mechanisms of action are not fully elucidated. In this research, we studied the anticancer potential of cisplatin at doses of 1, 2, or 3 µM using HL-60 cells as a test model. We investigated cisplatin effects at the molecular level using RNA sequencing, cell cycle analysis, and apoptotic assay after 24, 48, 72, and 96 hours of treatment. The results show that many genes responsible for molecular and cellular functions were significantly altered. Cisplatin treatment also caused the cells to be arrested at the DNA synthesis phase, and as the time increases, the cells gradually accumulated at the sub-G1 phase. Also, as the dose increases, a significant number of cells entered into the apoptotic and necrotic stages. Altogether, the data show that low doses of cisplatin significantly impact the viability of HL-60 cells, through modulation of gene expression, cell cycle, and apoptosis. PMID:27594783

  7. Histological alterations of intestinal villi and epithelial cells after feeding dietary sugar cane extract in piglets

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    Toshikazu Kawai

    2012-07-01

    Full Text Available Effects of sugar cane extract (SCE on the piglet intestinal histology were observed. Twelve castrated male piglets weaned at the age of 26 days were allotted to three groups fed diets containing 0, 0.05 or 0.10% SCE. At the end of feeding experiment, each intestinal segment was taken for light or scanning electron microscopy. Feed intake, body weight gain and feed efficiency did not show a difference among groups. Most of the values for villus height, villus area, cell area and cell mitosis numbers were not different among groups, except for that the villus area of the 0.10% SCE group and the cell area of both SCE groups increased significantly at the jejunum compared to the control (P<0.05. For cell mitosis numbers, the 0.10% SCE group was higher than the 0.05% SCE group at the jejunum. Compared with the majority of flat cells of each intestinal segment in the control, the SCE groups had protuberated cells. In the 0.05% SCE group, deeper cells at the sites of recently exfoliated cells in the duodenum, cell clusters aggregated by protuberated cells in the jejunum and much more protuberant cells in the ileum were observed. These histological intestinal alterations suggest that SCE could raise the functions of intestinal villi and epithelial cells, especially at the 0.05%.

  8. NK cells and type 1 innate lymphoid cells: partners in host defense.

    Science.gov (United States)

    Spits, Hergen; Bernink, Jochem H; Lanier, Lewis

    2016-06-21

    Innate lymphoid cells (ILCs) are effectors and regulators of innate immunity and tissue modeling and repair. Researchers have identified subsets of ILCs with differing functional activities, capacities to produce cytokines and transcription factors required for development and function. Natural killer (NK) cells represent the prototypical member of the ILC family. Together with ILC1s, NK cells constitute group 1 ILCs, which are characterized by their capacity to produce interferon-γ and their functional dependence on the transcription factor T-bet. NK cells and ILC1s are developmentally distinct but share so many features that they are difficult to distinguish, particularly under conditions of infection and inflammation. Here we review current knowledge of NK cells and the various ILC1 subsets. PMID:27328005

  9. Adipose tissue-derived mesenchymal stem cells as a new host cell in latent leishmaniasis.

    Science.gov (United States)

    Allahverdiyev, Adil M; Bagirova, Melahat; Elcicek, Serhat; Koc, Rabia Cakir; Baydar, Serap Yesilkir; Findikli, Necati; Oztel, Olga N

    2011-09-01

    Some protozoan infections such as Toxoplasma, Cryptosporidium, and Plasmodium can be transmitted through stem cell transplantations. To our knowledge, so far, there is no study about transmission of Leishmania parasites in stem cell transplantation and interactions between parasites and stem cells in vitro. Therefore, the aim of this study was to investigate the interaction between different species of Leishmania parasites and adipose tissue-derived mesenchymal stem cells (ADMSCs). ADMSCs have been isolated, cultured, characterized, and infected with different species of Leishmania parasites (L. donovani, L. major, L. tropica, and L. infantum). Infectivity was examined by Giemsa staining, microculture, and polymerase chain reaction methods. As a result, infectivity of ADMSCs by Leishmania parasites has been determined for the first time in this study. According to our findings, it is very important that donors are screened for Leishmania parasites before stem cell transplantations in regions where leishmaniasis is endemic. PMID:21896818

  10. Mechanistic Framework for Establishment, Maintenance, and Alteration of Cell Polarity in Plants

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    Pankaj Dhonukshe

    2012-01-01

    Full Text Available Cell polarity establishment, maintenance, and alteration are central to the developmental and response programs of nearly all organisms and are often implicated in abnormalities ranging from patterning defects to cancer. By residing at the distinct plasma membrane domains polar cargoes mark the identities of those domains, and execute localized functions. Polar cargoes are recruited to the specialized membrane domains by directional secretion and/or directional endocytic recycling. In plants, auxin efflux carrier PIN proteins display polar localizations in various cell types and play major roles in directional cell-to-cell transport of signaling molecule auxin that is vital for plant patterning and response programs. Recent advanced microscopy studies applied to single cells in intact plants reveal subcellular PIN dynamics. They uncover the PIN polarity generation mechanism and identified important roles of AGC kinases for polar PIN localization. AGC kinase family members PINOID, WAG1, and WAG2, belonging to the AGC-3 subclass predominantly influence the polar localization of PINs. The emerging mechanism for AGC-3 kinases action suggests that kinases phosphorylate PINs mainly at the plasma membrane after initial symmetric PIN secretion for eventual PIN internalization and PIN sorting into distinct ARF-GEF-regulated polar recycling pathways. Thus phosphorylation status directs PIN translocation to different cell sides. Based on these findings a mechanistic framework evolves that suggests existence of cell side-specific recycling pathways in plants and implicates AGC3 kinases for differential PIN recruitment among them for eventual PIN polarity establishment, maintenance, and alteration.

  11. Genetic barcode sequencing for screening altered population dynamics of hematopoietic stem cells transduced with lentivirus

    Science.gov (United States)

    Zanatta, Daniela B; Tsujita, Maristela; Borelli, Primavera; Aguiar, Rodrigo B; Ferrari, Daniel G; Strauss, Bryan E

    2014-01-01

    Insertional mutagenesis has been associated with malignant cell transformation in gene therapy protocols, leading to discussions about vector security. Therefore, clonal analysis is important for the assessment of vector safety and its impact on patient health. Here, we report a unique approach to assess dynamic changes in clonality of lentivirus transduced cells upon Sanger sequence analysis of a specially designed genetic barcode. In our approach, changes in the electropherogram peaks are measured and compared between successive time points, revealing alteration in the cell population. After in vitro validation, barcoded lentiviral libraries carrying IL2RG or LMO2 transgenes, or empty vector were used to transduce mouse hematopoietic (ckit+) stem cells, which were subsequently transplanted in recipient mice. We found that neither the empty nor IL2RG encoding vector had an effect on cell dynamics. In sharp contrast, the LMO2 oncogene was associated with altered cell dynamics even though hematologic counts remained unchanged, suggesting that the barcode could reveal changes in cell populations not observed by the frontline clinical assay. We describe a simple and sensitive method for the analysis of clonality, which could be easily used by any laboratory for the assessment of cellular behavior upon lentiviral transduction. PMID:26052520

  12. Altered features and increased chemosensitivity of human breast cancer cells mediated by adipose tissue-derived mesenchymal stromal cells

    International Nuclear Information System (INIS)

    Mesenchymal stromal cells (MSCs) represent heterogeneous cell population suitable for cell therapies in regenerative medicine. MSCs can also substantially affect tumor biology due to their ability to be recruited to the tumor stroma and interact with malignant cells via direct contacts and paracrine signaling. The aim of our study was to characterize molecular changes dictated by adipose tissue-derived mesenchymal stromal cells (AT-MSCs) and the effects on drug responses in human breast cancer cells SKBR3. The tumor cells were either directly cocultured with AT-MSCs or exposed to MSCs-conditioned medium (MSC-CM). Changes in cell biology were evaluated by kinetic live cell imaging, fluorescent microscopy, scratch wound assay, expression analysis, cytokine secretion profiling, ATP-based viability and apoptosis assays. The efficiency of cytotoxic treatment in the presence of AT-MSCs or MSCs-CM was analyzed. The AT-MSCs altered tumor cell morphology, induced epithelial-to-mesenchymal transition, increased mammosphere formation, cell confluence and migration of SKBR3. These features were attributed to molecular changes induced by MSCs-secreted cytokines and chemokines in breast cancer cells. AT-MSCs significantly inhibited the proliferation of SKBR3 cells in direct cocultures which was shown to be dependent on the SDF-1α/CXCR4 signaling axis. MSC-CM-exposed SKBR3 or SKBR3 in direct coculture with AT-MSCs exhibited increased chemosensitivity and induction of apoptosis in response to doxorubicin and 5-fluorouracil. Our work further highlights the multi-level nature of tumor-stromal cell interplay and demonstrates the capability of AT-MSCs and MSC-secreted factors to alter the anti-tumor drug responses

  13. CD133+ cells contribute to radioresistance via altered regulation of DNA repair genes in human lung cancer cells

    International Nuclear Information System (INIS)

    Background: Radioresistance in human tumors has been linked in part to a subset of cells termed cancer stem cells (CSCs). The prominin 1 (CD133) cell surface protein is proposed to be a marker enriching for CSCs. We explore the importance of DNA repair in contributing to radioresistance in CD133+ lung cancer cells. Materials and methods: A549 and H1299 lung cancer cell lines were used. Sorted CD133+ cells were exposed to either single 4 Gy or 8 Gy doses and clonogenic survival measured. ϒ-H2AX immunofluorescence and quantitative real time PCR was performed on sorted CD133+ cells both in the absence of IR and after two single 4 Gy doses. Lentiviral shRNA was used to silence repair genes. Results: A549 but not H1299 cells expand their CD133+ population after single 4 Gy exposure, and isolated A549 CD133+ cells demonstrate IR resistance. This resistance corresponded with enhanced repair of DNA double strand breaks (DSBs) and upregulated expression of DSB repair genes in A549 cells. Prior IR exposure of two single 4 Gy doses resulted in acquired DNA repair upregulation and improved repair proficiency in both A549 and H1299. Finally Exo1 and Rad51 silencing in A549 cells abrogated the CD133+ IR expansion phenotype and induced IR sensitivity in sorted CD133+ cells. Conclusions: CD133 identifies a population of cells within specific tumor types containing altered expression of DNA repair genes that are inducible upon exposure to chemotherapy. This altered gene expression contributes to enhanced DSB resolution and the radioresistance phenotype of these cells. We also identify DNA repair genes which may serve as promising therapeutic targets to confer radiosensitivity to CSCs

  14. Manipulation of host plant cells and tissues by gall-inducing insects and adaptive strategies used by different feeding guilds.

    Science.gov (United States)

    Oliveira, D C; Isaias, R M S; Fernandes, G W; Ferreira, B G; Carneiro, R G S; Fuzaro, L

    2016-01-01

    Biologists who study insect-induced plant galls are faced with the overwhelming diversity of plant forms and insect species. A challenge is to find common themes amidst this diversity. We discuss common themes that have emerged from our cytological and histochemical studies of diverse neotropical insect-induced galls. Gall initiation begins with recognition of reactive plant tissues by gall inducers, with subsequent feeding and/or oviposition triggering a cascade of events. Besides, to induce the gall structure insects have to synchronize their life cycle with plant host phenology. We predict that reactive oxygen species (ROS) play a role in gall induction, development and histochemical gradient formation. Controlled levels of ROS mediate the accumulation of (poly)phenols, and phytohormones (such as auxin) at gall sites, which contributes to the new cell developmental pathways and biochemical alterations that lead to gall formation. The classical idea of an insect-induced gall is a chamber lined with a nutritive tissue that is occupied by an insect that directly harvests nutrients from nutritive cells via its mouthparts, which function mechanically and/or as a delivery system for salivary secretions. By studying diverse gall-inducing insects we have discovered that insects with needle-like sucking mouthparts may also induce a nutritive tissue, whose nutrients are indirectly harvested as the gall-inducing insects feeds on adjacent vascular tissues. Activity of carbohydrate-related enzymes across diverse galls corroborates this hypothesis. Our research points to the importance of cytological and histochemical studies for elucidating mechanisms of induced susceptibility and induced resistance. PMID:26620152

  15. Manipulation of host plant cells and tissues by gall-inducing insects and adaptive strategies used by different feeding guilds.

    Science.gov (United States)

    Oliveira, D C; Isaias, R M S; Fernandes, G W; Ferreira, B G; Carneiro, R G S; Fuzaro, L

    2016-01-01

    Biologists who study insect-induced plant galls are faced with the overwhelming diversity of plant forms and insect species. A challenge is to find common themes amidst this diversity. We discuss common themes that have emerged from our cytological and histochemical studies of diverse neotropical insect-induced galls. Gall initiation begins with recognition of reactive plant tissues by gall inducers, with subsequent feeding and/or oviposition triggering a cascade of events. Besides, to induce the gall structure insects have to synchronize their life cycle with plant host phenology. We predict that reactive oxygen species (ROS) play a role in gall induction, development and histochemical gradient formation. Controlled levels of ROS mediate the accumulation of (poly)phenols, and phytohormones (such as auxin) at gall sites, which contributes to the new cell developmental pathways and biochemical alterations that lead to gall formation. The classical idea of an insect-induced gall is a chamber lined with a nutritive tissue that is occupied by an insect that directly harvests nutrients from nutritive cells via its mouthparts, which function mechanically and/or as a delivery system for salivary secretions. By studying diverse gall-inducing insects we have discovered that insects with needle-like sucking mouthparts may also induce a nutritive tissue, whose nutrients are indirectly harvested as the gall-inducing insects feeds on adjacent vascular tissues. Activity of carbohydrate-related enzymes across diverse galls corroborates this hypothesis. Our research points to the importance of cytological and histochemical studies for elucidating mechanisms of induced susceptibility and induced resistance.

  16. Impact of sub-inhibitory antibiotics on fibronectin-mediated host cell adhesion and invasion by Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Rasigade Jean

    2011-12-01

    Full Text Available Abstract Background Staphylococcus aureus is a well-armed pathogen prevalent in severe infections such as endocarditis and osteomyelitis. Fibronectin-binding proteins A and B, encoded by fnbA/B, are major pathogenesis determinants in these infections through their involvement in S. aureus adhesion to and invasion of host cells. Sub-minimum inhibitory concentrations (sub-MICs of antibiotics, frequently occurring in vivo because of impaired drug diffusion at the infection site, can alter S. aureus phenotype. We therefore investigated their impact on S. aureus fibronectin-mediated adhesiveness and invasiveness. Methods After in vitro challenge of S. aureus 8325-4 and clinical isolates with sub-MICs of major anti-staphylococcal agents, we explored fnbA/B transcription levels, bacterial adhesiveness to immobilised human fibronectin and human osteoblasts in culture, and bacterial invasion of human osteoblasts. Results Oxacillin, moxifloxacin and linezolid led to the development of a hyper-adhesive phenotype in the fibronectin adhesion assay that was consistent with an increase in fnbA/B transcription. Conversely, rifampin treatment decreased fibronectin binding in all strains tested without affecting fnbA/B transcription. Gentamicin and vancomycin had no impact on fibronectin binding or fnbA/B transcription levels. Only oxacillin-treated S. aureus displayed a significantly increased adhesion to cultured osteoblasts, but its invasiveness did not differ from that of untreated controls. Conclusion Our findings demonstrate that several antibiotics at sub-MICs modulate fibronectin binding in S. aureus in a drug-specific fashion. However, hyper- and hypo- adhesive phenotypes observed in controlled in vitro conditions were not fully confirmed in whole cell infection assays. The relevance of adhesion modulation during in vivo infections is thus still uncertain and requires further investigations.

  17. A novel method for producing target cells and assessing cytotoxic T lymphocyte activity in outbred hosts

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    Bendinelli Mauro

    2009-03-01

    Full Text Available Abstract Background Cytotoxic T lymphocytes play a crucial role in the immunological control of microbial infections and in the design of vaccines and immunotherapies. Measurement of cytotoxic T lymphocyte activity requires that the test antigen is presented by target cells having the same or compatible class I major hystocompatibility complex antigens as the effector cells. Conventional assays use target cells labeled with 51chromium and infer cytotoxic T lymphocyte activity by measuring the isotope released by the target cells lysed following incubation with antigen-specific cytotoxic T lymphocytes. This assay is sensitive but needs manipulation and disposal of hazardous radioactive reagents and provides a bulk estimate of the reporter released, which may be influenced by spontaneous release of the label and other poorly controllable variables. Here we describe a novel method for producing target in outbred hosts and assessing cytotoxic T lymphocyte activity by flow cytometry. Results The method consists of culturing skin fibroblasts, immortalizing them with a replication defective clone of simian virus 40, and finally transducing them with a bicistronic vector encoding the target antigen and the reporter green fluorescent protein. When used in a flow cytometry-based assay, the target cells obtained with this method proved valuable for assessing the viral envelope protein specific cytotoxic T lymphocyte activity in domestic cats acutely or chronically infected with feline immunodeficiency virus, a lentivirus similar to human immunodeficiency virus and used as animal model for AIDS studies. Conclusion Given the versatility of the bicistronic vector used, its ability to deliver multiple and large transgenes in target cells, and its extremely wide cell specificity when pseudotyped with the vesicular stomatitis virus envelope protein, the method is potentially exploitable in many animal species.

  18. Induction of Graft-versus-host Disease and In Vivo T Cell Monitoring Using an MHC-matched Murine Model

    OpenAIRE

    Anthony, Bryan A.; Hadley, Gregg A.

    2012-01-01

    Graft-versus-host disease (GVHD) is the limiting barrier to the broad use of bone marrow transplant as a curative therapy for a variety of hematological deficiencies. GVHD is caused by mature alloreactive T cells present in the bone marrow graft that are infused into the recipient and cause damage to host organs. However, in mice, T cells must be added to the bone marrow inoculum to cause GVHD. Although extensive work has been done to characterize T cell responses post transplant, bioluminesc...

  19. Harnessing the Effect of Adoptively Transferred Tumor-Reactive T Cells on Endogenous (Host-Derived Antitumor Immunity

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    Yolanda Nesbeth

    2010-01-01

    Full Text Available Adoptive T cell transfer therapy, the ex vivo activation, expansion, and subsequent administration of tumor-reactive T cells, is already the most effective therapy against certain types of cancer. However, recent evidence in animal models and clinical trials suggests that host conditioning interventions tailored for some of the most aggressive and frequent epithelial cancers will be needed to maximize the benefit of this approach. Similarly, the subsets, stage of differentiation, and ex vivo expansion procedure of tumor-reactive T cells to be adoptively transferred influence their in vivo effectiveness and may need to be adapted for different types of cancer and host conditioning interventions. The effects of adoptively transferred tumor-reactive T cells on the mechanisms of endogenous (host-derived antitumor immunity, and how to maximize their combined effects, are further discussed.

  20. Antigen presentation by murine epidermal langerhans cells and its alteration by ultraviolet B light

    International Nuclear Information System (INIS)

    Mice that are chronically exposed in vivo to ultraviolet B light (UV-B) display altered immunologic reactivity to various antigenic stimuli. A possible mode of UV-B action is that it exerts adverse effects on antigen-presenting cell function. Because the epidermis is the only tissue that is naturally subject to UV exposure we investigated if murine epidermal cells (EC) could perform an antigen presentation function and, if so, could this function be altered by UV-B irradiation. For this purpose, T cells immune to purified protein derivative of tuberculin (PPD) and dinitrophenylated ovalbumin (DNP6-OVA) from either BALB/c or C3H/He mice were incubated with syngeneic, semisyngeneic, or allogeneic EC or, for control purposes, with peritoneal exudate cells (PEC) that had been pulse-exposed to either the immunizing antigens or, as controls, left unpulsed, or pulsed to human serum albumin (HSA). After 4 days of culture, T cell proliferation was assessed by 3H-thymidine incorporation. PPD- and DNP/6-OVA pulsed, but not HSA-pulsed EC and PEC, induced vigorous proliferation of syngeneic and semisyngeneic, but not allogeneic, immune T cells. Pretreatment of stimulator cells with specific anti-Ia serum and complement virtually abolished this response, which indicated that among EC, Ia-bearing Langerhans cells are the critical stimulators. Exposure of EC either before or after pulsing to UV-B resulted in a dose-dependent impairment of antigen-specific T cell proliferation; the T proliferative response was abolished after administration of 20 mJ/cm2 UV-B. UV-B in the dose range employed did not produce immediate lethal cell damage, premature death of cultured EC, or toxic factors inhibitory for T cell proliferation

  1. WNT5A inhibits metastasis and alters splicing of Cd44 in breast cancer cells.

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    Wen Jiang

    Full Text Available Wnt5a is a non-canonical signaling Wnt. Low expression of WNT5A is correlated with poor prognosis in breast cancer patients. The highly invasive breast cancer cell lines, MDA-MB-231 and 4T1, express very low levels of WNT5A. To determine if enhanced expression of WNT5A would affect metastatic behavior, we generated WNT5A expressing cells from the 4T1 and MDA-MB-231 parental cell lines. WNT5A expressing cells demonstrated cobblestone morphology and reduced in vitro migration relative to controls. Cell growth was not altered. Metastasis to the lung via tail vein injection was reduced in the 4T1-WNT5A expressing cells relative to 4T1-vector controls. To determine the mechanism of WNT5A action on metastasis, we performed microarray and whole-transcriptome sequence analysis (RNA-seq to compare gene expression in 4T1-WNT5A and 4T1-vector cells. Analysis indicated highly significant alterations in expression of genes associated with cellular movement. Down-regulation of a subset of these genes, Mmp13, Nos2, Il1a, Cxcl2, and Lamb3, in WNT5A expressing cells was verified by semi-quantitative RT-PCR. Significant differences in transcript splicing were also detected in cell movement associated genes including Cd44. Cd44 is an adhesion molecule with a complex genome structure. Variable exon usage is associated with metastatic phenotype. Alternative spicing of Cd44 in WNT5A expressing cells was confirmed using RT-PCR. We conclude that WNT5A inhibits metastasis through down-regulation of multiple cell movement pathways by regulating transcript levels and splicing of key genes like Cd44.

  2. Csf2 Null Mutation Alters Placental Gene Expression and Trophoblast Glycogen Cell and Giant Cell Abundance in Mice1

    OpenAIRE

    Sferruzzi-Perri, Amanda N.; Macpherson, Anne M.; Roberts, Claire T.; Robertson, Sarah A.

    2009-01-01

    Genetic deficiency in granulocyte-macrophage colony-stimulating factor (CSF2, GM-CSF) results in altered placental structure in mice. To investigate the mechanism of action of CSF2 in placental morphogenesis, the placental gene expression and cell composition were examined in Csf2 null mutant and wild-type mice. Microarray and quantitative RT-PCR analyses on Embryonic Day (E) 13 placentae revealed that the Csf2 null mutation caused altered expression of 17 genes not previously known to be ass...

  3. An in vitro model of granuloma-like cell aggregates substantiates early host immune responses against Mycobacterium massiliense infection

    Science.gov (United States)

    Je, Sungmo; Quan, Hailian; Na, Yirang; Cho, Sang-Nae; Kim, Bum-Joon

    2016-01-01

    ABSTRACT Mycobacterium massiliense (M. mass), belonging to the M. abscessus complex, is a rapidly growing mycobacterium that is known to cause tuberculous-like lesions in humans. To better understand the interaction between host cells and M. mass, we used a recently developed in vitro model of early granuloma-like cell aggregates composed of human peripheral blood mononuclear cells (PBMCs). PBMCs formed granuloma-like, small and rounded cell aggregates when infected by live M. mass. Microscopic examination showed monocytes and macrophages surrounded by lymphocytes, which resembled cell aggregation induced by M. tuberculosis (M. tb). M. mass-infected PBMCs exhibited higher expression levels of HLA-DR, CD86 and CD80 on macrophages, and a significant decrease in the populations of CD4+ and CD8+ T cells. Interestingly, low doses of M. mass were sufficient to infect PBMCs, while active host cell death was gradually induced with highly increased bacterial loads, reflecting host destruction and dissemination of virulent rapid-growing mycobacteria (RGM). Collectively, this in vitro model of M. mass infection improves our understanding of the interplay of host immune cells with mycobacteria, and may be useful for developing therapeutics to control bacterial pathogenesis. PMID:27489303

  4. Inherent Variability of Growth Media Impacts the Ability of Salmonella Typhimurium to Interact with Host Cells.

    Science.gov (United States)

    Sridhar, Sushmita; Steele-Mortimer, Olivia

    2016-01-01

    Efficient invasion of non-phagocytic cells, such as intestinal epithelial cells, by Salmonella Typhimurium is dependent on the Salmonella Pathogenicity Island 1 (SPI-1)-encoded Type Three Secretion System. The environmental cues involved in SPI-1 induction are not well understood. In vitro, various conditions are used to induce SPI-1 and the invasive phenotype. Although lysogeny broth (LB) is widely used, multiple formulations exist, and variation can arise due to intrinsic differences in complex components. Minimal media are also susceptible to variation. Still, the impact of these inconsistencies on Salmonella virulence gene expression has not been well studied. The goal of this project is to identify growth conditions in LB and minimal medium that affect SPI-1 induction in vitro using both whole population and single cell analysis. Here we show, using a fluorescent reporter of the SPI-1 gene prgH, that growth of Salmonella in LB yields variable induction. Deliberate modification of media components can influence the invasive profile. Finally, we demonstrate that changes in SPI-1 inducing conditions can affect the ability of Salmonella to replicate intracellularly. These data indicate that the specific media growth conditions impact how the bacteria interact with host cells.

  5. Extracellular Vesicle-Associated Transitory Cell Wall Components and Their Impact on the Interaction of Fungi with Host Cells.

    Science.gov (United States)

    Nimrichter, Leonardo; de Souza, Marcio M; Del Poeta, Maurizio; Nosanchuk, Joshua D; Joffe, Luna; Tavares, Patricia de M; Rodrigues, Marcio L

    2016-01-01

    Classic cell wall components of fungi comprise the polysaccharides glucans and chitin, in association with glycoproteins and pigments. During the last decade, however, system biology approaches clearly demonstrated that the composition of fungal cell walls include atypical molecules historically associated with intracellular or membrane locations. Elucidation of mechanisms by which many fungal molecules are exported to the extracellular space suggested that these atypical components are transitorily located to the cell wall. The presence of extracellular vesicles (EVs) at the fungal cell wall and in culture supernatants of distinct pathogenic species suggested a highly functional mechanism of molecular export in these organisms. Thus, the passage of EVs through fungal cell walls suggests remarkable molecular diversity and, consequently, a potentially variable influence on the host antifungal response. On the basis of information derived from the proteomic characterization of fungal EVs from the yeasts Cryptoccocus neoformans and Candida albicans and the dimorphic fungi Histoplasma capsulatum and Paracoccidioides brasiliensis, our manuscript is focused on the clear view that the fungal cell wall is much more complex than previously thought. PMID:27458437

  6. Extracellular Vesicle-Associated Transitory Cell Wall Components and Their Impact on the Interaction of Fungi with Host Cells

    Science.gov (United States)

    Nimrichter, Leonardo; de Souza, Marcio M.; Del Poeta, Maurizio; Nosanchuk, Joshua D.; Joffe, Luna; Tavares, Patricia de M.; Rodrigues, Marcio L.

    2016-01-01

    Classic cell wall components of fungi comprise the polysaccharides glucans and chitin, in association with glycoproteins and pigments. During the last decade, however, system biology approaches clearly demonstrated that the composition of fungal cell walls include atypical molecules historically associated with intracellular or membrane locations. Elucidation of mechanisms by which many fungal molecules are exported to the extracellular space suggested that these atypical components are transitorily located to the cell wall. The presence of extracellular vesicles (EVs) at the fungal cell wall and in culture supernatants of distinct pathogenic species suggested a highly functional mechanism of molecular export in these organisms. Thus, the passage of EVs through fungal cell walls suggests remarkable molecular diversity and, consequently, a potentially variable influence on the host antifungal response. On the basis of information derived from the proteomic characterization of fungal EVs from the yeasts Cryptoccocus neoformans and Candida albicans and the dimorphic fungi Histoplasma capsulatum and Paracoccidioides brasiliensis, our manuscript is focused on the clear view that the fungal cell wall is much more complex than previously thought. PMID:27458437

  7. Apoptotic-like Leishmania exploit the host´s autophagy machinery to reduce T-cell-mediated parasite elimination

    Science.gov (United States)

    Crauwels, Peter; Bohn, Rebecca; Thomas, Meike; Gottwalt, Stefan; Jäckel, Florian; Krämer, Susi; Bank, Elena; Tenzer, Stefan; Walther, Paul; Bastian, Max; van Zandbergen, Ger

    2015-01-01

    Apoptosis is a well-defined cellular process in which a cell dies, characterized by cell shrinkage and DNA fragmentation. In parasites like Leishmania, the process of apoptosis-like cell death has been described. Moreover upon infection, the apoptotic-like population is essential for disease development, in part by silencing host phagocytes. Nevertheless, the exact mechanism of how apoptosis in unicellular organisms may support infectivity remains unclear. Therefore we investigated the fate of apoptotic-like Leishmania parasites in human host macrophages. Our data showed—in contrast to viable parasites—that apoptotic-like parasites enter an LC3+, autophagy-like compartment. The compartment was found to consist of a single lipid bilayer, typical for LC3-associated phagocytosis (LAP). As LAP can provoke anti-inflammatory responses and autophagy modulates antigen presentation, we analyzed how the presence of apoptotic-like parasites affected the adaptive immune response. Macrophages infected with viable Leishmania induced proliferation of CD4+ T-cells, leading to a reduced intracellular parasite survival. Remarkably, the presence of apoptotic-like parasites in the inoculum significantly reduced T-cell proliferation. Chemical induction of autophagy in human monocyte-derived macrophage (hMDM), infected with viable parasites only, had an even stronger proliferation-reducing effect, indicating that host cell autophagy and not parasite viability limits the T-cell response and enhances parasite survival. Concluding, our data suggest that apoptotic-like Leishmania hijack the host cells´ autophagy machinery to reduce T-cell proliferation. Furthermore, the overall population survival is guaranteed, explaining the benefit of apoptosis-like cell death in a single-celled parasite and defining the host autophagy pathway as a potential therapeutic target in treating Leishmaniasis. PMID:25801301

  8. Stem cell recruitment of newly formed host cells via a successful seduction? Filling the gap between neurogenic niche and injured brain site.

    Directory of Open Access Journals (Sweden)

    Naoki Tajiri

    Full Text Available Here, we report that a unique mechanism of action exerted by stem cells in the repair of the traumatically injured brain involves their ability to harness a biobridge between neurogenic niche and injured brain site. This biobridge, visualized immunohistochemically and laser captured, corresponded to an area between the neurogenic subventricular zone and the injured cortex. That the biobridge expressed high levels of extracellular matrix metalloproteinases characterized initially by a stream of transplanted stem cells, but subsequently contained only few to non-detectable grafts and overgrown by newly formed host cells, implicates a novel property of stem cells. The transplanted stem cells manifest themselves as pathways for trafficking the migration of host neurogenic cells, but once this biobridge is formed between the neurogenic site and the injured brain site, the grafted cells disappear and relinquish their task to the host neurogenic cells. Our findings reveal that long-distance migration of host cells from the neurogenic niche to the injured brain site can be achieved through transplanted stem cells serving as biobridges for initiation of endogenous repair mechanisms. This is the first report of a stem cell-paved "biobridge". Indeed, to date the two major schools of discipline in stem cell repair mechanism primarily support the concept of "cell replacement" and bystander effects of "trophic factor secretion". The present novel observations of a stem cell seducing a host cell to engage in brain repair advances basic science concepts on stem cell biology and extracellular matrix, as well as provokes translational research on propagating this stem cell-paved biobridge beyond cell replacement and trophic factor secretion for the treatment of traumatic brain injury and other neurological disorders.

  9. Stem cell recruitment of newly formed host cells via a successful seduction? Filling the gap between neurogenic niche and injured brain site.

    Science.gov (United States)

    Tajiri, Naoki; Kaneko, Yuji; Shinozuka, Kazutaka; Ishikawa, Hiroto; Yankee, Ernest; McGrogan, Michael; Case, Casey; Borlongan, Cesar V

    2013-01-01

    Here, we report that a unique mechanism of action exerted by stem cells in the repair of the traumatically injured brain involves their ability to harness a biobridge between neurogenic niche and injured brain site. This biobridge, visualized immunohistochemically and laser captured, corresponded to an area between the neurogenic subventricular zone and the injured cortex. That the biobridge expressed high levels of extracellular matrix metalloproteinases characterized initially by a stream of transplanted stem cells, but subsequently contained only few to non-detectable grafts and overgrown by newly formed host cells, implicates a novel property of stem cells. The transplanted stem cells manifest themselves as pathways for trafficking the migration of host neurogenic cells, but once this biobridge is formed between the neurogenic site and the injured brain site, the grafted cells disappear and relinquish their task to the host neurogenic cells. Our findings reveal that long-distance migration of host cells from the neurogenic niche to the injured brain site can be achieved through transplanted stem cells serving as biobridges for initiation of endogenous repair mechanisms. This is the first report of a stem cell-paved "biobridge". Indeed, to date the two major schools of discipline in stem cell repair mechanism primarily support the concept of "cell replacement" and bystander effects of "trophic factor secretion". The present novel observations of a stem cell seducing a host cell to engage in brain repair advances basic science concepts on stem cell biology and extracellular matrix, as well as provokes translational research on propagating this stem cell-paved biobridge beyond cell replacement and trophic factor secretion for the treatment of traumatic brain injury and other neurological disorders.

  10. Induction of Central Host Signaling Kinases during Pneumococcal Infection of Human THP-1 Cells.

    Science.gov (United States)

    Kohler, Thomas P; Scholz, Annemarie; Kiachludis, Delia; Hammerschmidt, Sven

    2016-01-01

    bacterial uptake. The participation of essential host cell signaling kinases in pneumococcal phagocytosis was deciphered for the kinase Akt, ERK1/2, and p38 and phosphoimmunoblots showed an increased phosphorylation and thus activation upon infection with pneumococci. Taken together, this study deciphers host cell kinases in innate immune cells that are induced upon infection with pneumococci and interfere with bacterial clearance after phagocytosis. PMID:27200303

  11. Induction of central host signalling kinases during pneumococcal infection of human THP-1 cells

    Directory of Open Access Journals (Sweden)

    Thomas Peter Kohler

    2016-04-01

    as well during bacterial uptake. The participation of essential host cell signaling kinases in pneumococcal phagocytosis was deciphered for the kinase Akt, ERK1/2 and p38 and phosphoimmunoblots showed an increased phosphorylation and thus activation upon infection with pneumococci. Taken together, this study deciphers host cell kinases in innate immune cells that are induced upon infection with pneumococci and interfere with bacterial clearance after phagocytosis.

  12. MicroRNA and DNA methylation alterations mediating retinoic acid induced neuroblastoma cell differentiation.

    Science.gov (United States)

    Stallings, Raymond L; Foley, Niamh H; Bray, Isabella M; Das, Sudipto; Buckley, Patrick G

    2011-10-01

    Many neuroblastoma cell lines can be induced to differentiate into a mature neuronal cell type with retinoic acid and other compounds, providing an important model system for elucidating signalling pathways involved in this highly complex process. Recently, it has become apparent that miRNAs, which act as regulators of gene expression at a post-transcriptional level, are differentially expressed in differentiating cells and play important roles governing many aspects of this process. This includes the down-regulation of DNA methyltransferases that cause the de-methylation and transcriptional activation of numerous protein coding gene sequences. The purpose of this article is to review involvement of miRNAs and DNA methylation alterations in the process of neuroblastoma cell differentiation. A thorough understanding of miRNA and genetic pathways regulating neuroblastoma cell differentiation potentially could lead to targeted therapies for this disease.

  13. Systematics of hydrothermal alteration at the volcanic-hosted Falun Zn-Pb-Cu-(Au-Ag) deposit - implications for ore genesis, structure and exploration in a 1.9 Ga ore district, Fennoscandian Shield, Sweden

    Science.gov (United States)

    Kampmann, Tobias C.; Jansson, Nils J.; Stephens, Michael B.; Majka, Jarosław

    2016-04-01

    The Palaeoproterozoic, volcanic-hosted Falun Zn-Pb-Cu-(Au-Ag) sulphide deposit was mined for base and precious metals during several centuries, until its closure in 1992. The deposit is located in a 1.9 Ga ore district in the Bergslagen lithotectonic unit, Fennoscandian Shield, south-central Sweden. Both the ores and their host rock underwent polyphase ductile deformation, and metamorphism under amphibolite facies and later retrograde conditions at 1.9-1.8 Ga (Svecokarelian orogenic system). This study has the following aims: (i) Classify styles and intensities of alteration in the hydrothermally altered zone at Falun; (ii) identify precursor rocks to hydrothermally altered rocks and their spatial distribution at the deposit; (iii) evaluate the chemical changes resulting from hydrothermal alteration using mass change calculations; and (iv) assess the pre-metamorphic alteration assemblages accounting for the observed metamorphic mineral associations in the altered rocks at Falun. Results will have implications for both the ore-genetic and structural understanding of the deposit, as well as for local and regional exploration. Metamorphic mineral associations in the altered rocks include biotite-quartz-cordierite-(anthophyllite) and, more proximally, quartz-anthophyllite-(biotite-cordierite/almandine), biotite-cordierite-(anthophyllite) and biotite-almandine-(anthophyllite). The proximal hydrothermally altered zone corresponds to intense chlorite-style alteration. Subordinate dolomite or calcite marble, as well as calc-silicate (tremolite, diopside) rocks are also present at the deposit. Metavolcanic rocks around the deposit are unaltered, weakly sericitized or sodic-altered. Immobile-element (e.g. Zr, TiO2, Al2O3, REE) systematics of the silicate-rich samples at and around the deposit suggest that the precursors to the hydrothermally altered rocks at Falun were predominantly rhyolitic in composition, dacitic rocks being subordinate and mafic-intermediate rocks

  14. Chromosomal and Nuclear Alterations in Root Tip Cells of Allium Cepa L. Induced by Alprazolam

    Science.gov (United States)

    Nefic, Hilada; Musanovic, Jasmin; Metovic, Azra; Kurteshi, Kemajl

    2013-01-01

    ABSTRACT Introduction: Alprazolam is a triazolobenzodiazepine used in panic disorders and other anxiety states. Target organ of Alprazolam is CNS, causing depression of respiration and consciousness. Aim: This study aimed to estimate the genotoxic potential of Alprazolam using Allium cepa test. Methods: Allium cepa is one of the most suitable plants for detecting different types of xenobiotics. The test enables the assessment of different genetic endpoints making possible damage to the DNA of humans to be predicted. Results: Alprazolam induced chromosomal (anaphase bridges, breaks, lagging and stickiness, abnormal spiralisation, multipolarity and polyploidy) and cytological aberrations, especially nuclear alterations (nuclear buds, fragmented nucleus and apoptotic bodies, cells without nucleus, binucleated and micronucleated cells), morphological alterations in shape and size of cells, spindle disturbance and polar deviation in root tip meristem cells of Allium cepa at all tested concentrations. Alprazolam also caused significant inhibition of mitotic index in these cells. Conclusion: These changes in cells are indicators of genotoxic potential of Alprazolam suggesting a need for further in vitro studies on animal and human lymphocytes as well as in vivo studies. PMID:25568504

  15. Mycobacterium tuberculosis nuoG is a virulence gene that inhibits apoptosis of infected host cells.

    Directory of Open Access Journals (Sweden)

    Kamalakannan Velmurugan

    2007-07-01

    Full Text Available The survival and persistence of Mycobacterium tuberculosis depends on its capacity to manipulate multiple host defense pathways, including the ability to actively inhibit the death by apoptosis of infected host cells. The genetic basis for this anti-apoptotic activity and its implication for mycobacterial virulence have not been demonstrated or elucidated. Using a novel gain-of-function genetic screen, we demonstrated that inhibition of infection-induced apoptosis of macrophages is controlled by multiple genetic loci in M. tuberculosis. Characterization of one of these loci in detail revealed that the anti-apoptosis activity was attributable to the type I NADH-dehydrogenase of M. tuberculosis, and was mainly due to the subunit of this multicomponent complex encoded by the nuoG gene. Expression of M. tuberculosis nuoG in nonpathogenic mycobacteria endowed them with the ability to inhibit apoptosis of infected human or mouse macrophages, and increased their virulence in a SCID mouse model. Conversely, deletion of nuoG in M. tuberculosis ablated its ability to inhibit macrophage apoptosis and significantly reduced its virulence in mice. These results identify a key component of the genetic basis for an important virulence trait of M. tuberculosis and support a direct causal relationship between virulence of pathogenic mycobacteria and their ability to inhibit macrophage apoptosis.

  16. Roles of the Picornaviral 3C Proteinase in the Viral Life Cycle and Host Cells

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    Di Sun

    2016-03-01

    Full Text Available The Picornaviridae family comprises a large group of non-enveloped viruses that have a major impact on human and veterinary health. The viral genome contains one open reading frame encoding a single polyprotein that can be processed by viral proteinases. The crucial 3C proteinases (3Cpros of picornaviruses share similar spatial structures and it is becoming apparent that 3Cpro plays a significant role in the viral life cycle and virus host interaction. Importantly, the proteinase and RNA-binding activity of 3Cpro are involved in viral polyprotein processing and the initiation of viral RNA synthesis. In addition, 3Cpro can induce the cleavage of certain cellular factors required for transcription, translation and nucleocytoplasmic trafficking to modulate cell physiology for viral replication. Due to interactions between 3Cpro and these essential factors, 3Cpro is also involved in viral pathogenesis to support efficient infection. Furthermore, based on the structural conservation, the development of irreversible inhibitors and discovery of non-covalent inhibitors for 3Cpro are ongoing and a better understanding of the roles played by 3Cpro may provide insights into the development of potential antiviral treatments. In this review, the current knowledge regarding the structural features, multiple functions in the viral life cycle, pathogen host interaction, and development of antiviral compounds for 3Cpro is summarized.

  17. Direct identification of the Meloidogyne incognita secretome reveals proteins with host cell reprogramming potential.

    Directory of Open Access Journals (Sweden)

    Stéphane Bellafiore

    2008-10-01

    Full Text Available The root knot nematode, Meloidogyne incognita, is an obligate parasite that causes significant damage to a broad range of host plants. Infection is associated with secretion of proteins surrounded by proliferating cells. Many parasites are known to secrete effectors that interfere with plant innate immunity, enabling infection to occur; they can also release pathogen-associated molecular patterns (PAMPs, e.g., flagellin that trigger basal immunity through the nematode stylet into the plant cell. This leads to suppression of innate immunity and reprogramming of plant cells to form a feeding structure containing multinucleate giant cells. Effectors have generally been discovered using genetics or bioinformatics, but M. incognita is non-sexual and its genome sequence has not yet been reported. To partially overcome these limitations, we have used mass spectrometry to directly identify 486 proteins secreted by M. incognita. These proteins contain at least segmental sequence identity to those found in our 3 reference databases (published nematode proteins; unpublished M. incognita ESTs; published plant proteins. Several secreted proteins are homologous to plant proteins, which they may mimic, and they contain domains that suggest known effector functions (e.g., regulating the plant cell cycle or growth. Others have regulatory domains that could reprogram cells. Using in situ hybridization we observed that most secreted proteins were produced by the subventral glands, but we found that phasmids also secreted proteins. We annotated the functions of the secreted proteins and classified them according to roles they may play in the development of root knot disease. Our results show that parasite secretomes can be partially characterized without cognate genomic DNA sequence. We observed that the M. incognita secretome overlaps the reported secretome of mammalian parasitic nematodes (e.g., Brugia malayi, suggesting a common parasitic behavior and a possible

  18. Altered microRNAs expression profiling in cumulus cells from patients with polycystic ovary syndrome

    OpenAIRE

    Liu, Suying; Zhang, Xuan; Shi, Changgen; Lin, Jimin; Chen, Guowu; Wu, Bin; Wu, Ligang; Shi, Huijuan; Yuan, Yao; Zhou, Weijin; Sun, Zhaogui; Dong, Xi; Wang, Jian

    2015-01-01

    Background Polycystic ovary syndrome (PCOS) is a common endocrine disorder in women of reproductive age, and oocyte developmental competence is altered in patients with PCOS. In recent years microRNAs (miRNAs) have emerged as important regulators of gene expression, the aim of the study was to study miRNAs expression patterns of cumulus cells from PCOS patients. Methods The study included 20 patients undergoing in vitro fertilization (IVF) and intra-cytoplasmic sperm injection (ICSI): 10 diag...

  19. Targeted alteration of real and imaginary refractive index of biological cells by histological staining

    OpenAIRE

    Cherkezyan, Lusik; Subramanian, Hariharan; Stoyneva, Valentina; Rogers, Jeremy D.; Yang, Seungmoo; Damania, Dhwanil; Taflove, Allen; Backman, Vadim

    2012-01-01

    Various staining techniques are commonly used in biomedical research to investigate cellular morphology. By inducing absorption of light, staining dyes change the intracellular refractive index due to the Kramers-Kronig relationship. We present a method for creating 2-D maps of real and imaginary refractive indices of stained biological cells using their thickness and absorptance. We validate our technique on dyed polystyrene microspheres and quantify the alteration in refractive index of sta...

  20. Alterations in kainate receptor and TRPM1 localization in bipolar cells after retinal photoreceptor degeneration

    Directory of Open Access Journals (Sweden)

    Jacqueline eGayet-Primo

    2015-12-01

    Full Text Available Photoreceptor degeneration differentially impacts glutamatergic signaling in downstream On and Off bipolar cells. In rodent models, photoreceptor degeneration leads to loss of glutamatergic signaling in On bipolar cells, whereas Off bipolar cells appear to retain glutamate sensitivity, even after extensive photoreceptor loss. The localization and identity of the receptors that mediate these residual glutamate responses in Off bipolar cells have not been determined. Recent studies show that macaque and mouse Off bipolar cells receive glutamatergic input primarily through kainate-type glutamate receptors. Here, we studied the impact of photoreceptor degeneration on glutamate receptor associated proteins in Off and On bipolar cells. We show that the kainate receptor subunit, GluK1, persists in remodeled Off bipolar cell dendrites of the rd10 mouse retina. However, the pattern of expression is altered and the intensity of staining is reduced compared to wild-type retina. The kainate receptor auxiliary subunit, Neto1, also remains in Off bipolar cell dendrites after complete photoreceptor degeneration. Similar preservation of kainate receptor subunits was evident in human retina in which photoreceptors had degenerated due to serous retinal detachment. In contrast, photoreceptor degeneration leads to loss of synaptic expression of TRPM1 in mouse and human On bipolar cells, but strong somatic expression remains. These findings demonstrate that Off bipolar cells retain dendritic glutamate receptors during retinal degeneration and could thus serve as a conduit for signal transmission from transplanted or optogenetically-restored photoreceptors.

  1. Alteration of pancreatic cancer cell functions by tumor-stromal cell interaction

    OpenAIRE

    Shin eHamada; Atsushi eMasamune; Tooru eShimosegawa

    2013-01-01

    Pancreatic cancer shows a characteristic tissue structure called desmoplasia, which consists of dense fibrotic stroma surrounding cancer cells. Interactions between pancreatic cancer cells and stromal cells promote invasive growth of cancer cells and establish a specific microenvironment such as hypoxia which further aggravates the malignant behavior of cancer cells. Pancreatic stellate cells (PSCs) play pivotal role in the development of fibrosis within the pancreatic cancer tissue, and also...

  2. Alteration of pancreatic cancer cell functions by tumor-stromal cell interaction

    OpenAIRE

    Hamada, Shin; Masamune, Atsushi; Shimosegawa, Tooru

    2013-01-01

    Pancreatic cancer shows a characteristic tissue structure called desmoplasia, which consists of dense fibrotic stroma surrounding cancer cells. Interactions between pancreatic cancer cells and stromal cells promote invasive growth of cancer cells and establish a specific microenvironment such as hypoxia which further aggravates the malignant behavior of cancer cells. Pancreatic stellate cells (PSCs) play a pivotal role in the development of fibrosis within the pancreatic cancer tissue, and al...

  3. Menstrual cycle distribution of uterine natural killer cells is altered in heavy menstrual bleeding.

    Science.gov (United States)

    Biswas Shivhare, Sourima; Bulmer, Judith N; Innes, Barbara A; Hapangama, Dharani K; Lash, Gendie E

    2015-11-01

    Heavy menstrual bleeding (HMB) affects 30% of women of reproductive age and significantly interferes with quality of life. Altered endometrial vascular maturation has been reported in HMB and recurrent miscarriage, the latter associated with increased uterine natural killer (uNK) cell numbers. This study compared endometrial leukocyte populations in controls and women with HMB. Formalin-fixed paraffin-embedded endometrial biopsies from controls (without endometrial pathology) and HMB were immunostained for CD14 (macrophages), CD56 (uNK cells), CD83 (dendritic cells), FOXP3 (regulatory T cells/Tregs), CD3 and CD8 (T cells). Leukocyte numbers were analysed as a percentage of total stromal cells in five randomly selected fields of view in the stratum functionalis of each sample. In control women across the menstrual cycle, 2-8% of total stromal cells were CD3(+) cells, 2-4% were CD8(+) T cells and 6-8% were CD14(+) macrophages. Compared with controls, CD3(+) cells were reduced during the mid-secretory phase (4%, P<0.01) and increased in the late secretory phase (12%, P=0.01) in HMB. CD83(+) dendritic cells and FOXP3(+) Tregs were scarce throughout the menstrual cycle in both groups. In controls, 2% of stromal cells in proliferative endometrium were CD56(+) uNK cells, increasing to 17% during the late secretory phase. In HMB, CD56(+) uNK cells were increased in the proliferative (5%, P<0.01) and early secretory (4%, P<0.02) phases, but reduced (10%, P<0.01) in the late secretory phase. This study demonstrates dysregulation of uNK cells in HMB, the functional consequence of which may have an impact on endometrial vascular development and/or endometrial preparation for menstruation.

  4. HCMV-infected cells maintain efficient nucleotide excision repair of the viral genome while abrogating repair of the host genome.

    Directory of Open Access Journals (Sweden)

    John M O'Dowd

    Full Text Available Many viruses subvert the host cell's ability to mount and complete various DNA damage responses (DDRs after infection. HCMV infection of permissive fibroblasts activates host DDRs at the time of viral deposition and during replication, but the DDRs remain uncompleted without arrest or apoptosis. We believe this was in part due to partitioning of the damage response and double strand break repair components. After extraction of soluble proteins, the localization of these components fell into three groups: specifically associated with the viral replication centers (RCs, diffused throughout the nucleoplasm and excluded from the RCs. Others have shown that cells are incapable of processing exogenously introduced damage after infection. We hypothesized that the inability of the cells to process damage might be due to the differential association of repair components within the RCs and, in turn, potentially preferential repair of the viral genome and compromised repair of the host genome. To test this hypothesis we used multiple strategies to examine repair of UV-induced DNA damage in mock and virus-infected fibroblasts. Comet assays indicated that repair was initiated, but was not completed in infected cells. Quantitative analysis of immunofluorescent localization of cyclobutane pyrimidine dimers (CPDs revealed that after 24 h of repair, CPDs were significantly reduced in viral DNA, but not significantly changed in the infected host DNA. To further quantitate CPD repair, we developed a novel dual-color Southern protocol allowing visualization of host and viral DNA simultaneously. Combining this Southern methodology with a CPD-specific T4 endonuclease V alkaline agarose assay to quantitate repair of adducts, we found efficient repair of CPDs from the viral DNA but not host cellular DNA. Our data confirm that NER functions in HCMV-infected cells and almost exclusively repairs the viral genome to the detriment of the host's genome.

  5. Analysis of Alterations in Morphologic Characteristics of Mesenchymal Stem Cells by Mechanical Stimulation during Differentiation into Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Mohammad Ali Shokrgozar

    2010-01-01

    Full Text Available Objective: Mesenchymal stem cells (MSCs can be expanded and differentiated intomany mature cell types including smooth muscle cells (SMCs. In addition to growth factor,cyclic stretch contributes to differentiation of stem cells. Mechanical stimuli are criticalto morphological changes, development, regeneration, differentiation and pathology ofmesenchymal tissues. The aim of this study is to investigate effects of cyclic stretch withdiffering amplitudes on morphology and differentiation of mesenchymal stem cells.Materials and Methods: Mesenchymal stem cells are extracted from human bone marrow.Cells are cultured on silicone membrane and exposed to cyclic stretch by a custommade device. Cellular images are captured before and after tests. Effects of 5% and 15%uniaxial strain with 1Hz frequency and 1-8 hour durations on morphology of human mesenchymalstem cells are investigated. It is assumed that environmental factors such asmechanical loading regulate MSCs differentiation to SMCs. Fractal analysis is used toquantify alterations in cellular morphology. An image processing method with a designedcode is used for evaluation of fractal dimension parameter.Results: Results demonstrate statistically significant change in cell morphology due tomechanical stretch. By elevation of strain amplitude and number of load cycles, fractaldimensions of cell images decrease. Such decrease is equivalent to alignment of cells bymechanical stimulus. Cells are differentiated to SMCs purely by cyclic stretch. The initiationand rate of differentiation depend on mechanical conditions.Conclusion: To produce functional SMCs for engineered tissues, MSCs can be exposed to uniaxialcyclic stretch. The functionality of differentiated SMCs depends on loading conditions.

  6. Loss of caveolin-1 causes blood-retinal barrier breakdown, venous enlargement, and mural cell alteration.

    Science.gov (United States)

    Gu, Xiaowu; Fliesler, Steven J; Zhao, You-Yang; Stallcup, William B; Cohen, Alex W; Elliott, Michael H

    2014-02-01

    Blood-retinal barrier (BRB) breakdown and related vascular changes are implicated in several ocular diseases. The molecules and mechanisms regulating BRB integrity and pathophysiology are not fully elucidated. Caveolin-1 (Cav-1) ablation results in loss of caveolae and microvascular pathologies, but the role of Cav-1 in the retina is largely unknown. We examined BRB integrity and vasculature in Cav-1 knockout mice and found a significant increase in BRB permeability, compared with wild-type controls, with branch veins being frequent sites of breakdown. Vascular hyperpermeability occurred without apparent alteration in junctional proteins. Such hyperpermeability was not rescued by inhibiting eNOS activity. Veins of Cav-1 knockout retinas exhibited additional pathological features, including i) eNOS-independent enlargement, ii) altered expression of mural cell markers (eg, down-regulation of NG2 and up-regulation of αSMA), and iii) dramatic alterations in mural cell phenotype near the optic nerve head. We observed a significant NO-dependent increase in retinal artery diameter in Cav-1 knockout mice, suggesting that Cav-1 plays a role in autoregulation of resistance vessels in the retina. These findings implicate Cav-1 in maintaining BRB integrity in retinal vasculature and suggest a previously undefined role in the retinal venous system and associated mural cells. Our results are relevant to clinically significant retinal disorders with vascular pathologies, including diabetic retinopathy, uveoretinitis, and primary open-angle glaucoma.

  7. Cell surface proteome analysis of human-hosted Trypanosoma cruzi life stages

    DEFF Research Database (Denmark)

    Queiroz, Rayner M L; Charneau, Sébastien; Bastos, Izabela M D;

    2014-01-01

    addressed the analysis of the plasma membrane (PM) subproteome from T. cruzi human-hosted life stages, trypomastigote and axenic amastigote, by two complementary PM protein enrichment techniques followed by identification using an LC-MS/MS approach. The results revealed an extensive repertoire of proteins...... in the PM subproteomes, including enzymes that might be suitable candidates for drug intervention. The comparison of the cell surface proteome among the life forms revealed some potentially stage-specific enzymes, although the majority was shared by both stages. Bioinformatic analysis showed that the vast......Chagas' disease is a neglected infectious illness, caused by the protozoan Trypanosoma cruzi. It remains a challenging health issue in Latin America, where it is endemic, and so far there is no immunoprophylatic vaccine or satisfactory chemotherapic treatment for its chronic stage. The present work...

  8. Interleukin 2 expression by tumor cells alters both the immune response and the tumor microenvironment.

    Science.gov (United States)

    Lee, J; Fenton, B M; Koch, C J; Frelinger, J G; Lord, E M

    1998-04-01

    Microenvironmental conditions within solid tumors can have marked effects on the growth of the tumors and their response to therapies. The disorganized growth of tumors and their attendant vascular systems tends to result in areas of the tumors that are deficient in oxygen (hypoxic). Cells within these hypoxic areas are more resistant to conventional therapies such as radiation and chemotherapy. Here, we examine the hypoxic state of EMT6 mouse mammary tumors and the location of host cells within the different areas of the tumors to determine whether such microenvironmental conditions might also affect their ability to be recognized by the immune system. Hypoxia within tumors was quantified by flow cytometry and visualized by immunohistochemistry using a monoclonal antibody (ELK3-51) against cellular adducts of 2-(2-nitro-1H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetam ide (EF5), a nitroimidazole compound that binds selectively to hypoxic cells. Thy-1+ cells, quantified using a monoclonal antibody, were found only in the well-oxygenated areas. The location of these Thy-1+ cells was also examined in EMT6 tumors that had been transfected with the gene for interleukin-2 (IL-2) because these tumors contain greatly increased numbers of host cells. Surprisingly, we found that IL-2-transfected tumors had significantly decreased hypoxia compared to parental tumors. Furthermore, using the fluorescent dye Hoechst 33342, an in vivo marker of perfused vessels, combined with immunochemical staining of PECAM-1 (CD31) as a marker of tumor vasculature, we found increased vascularization in the IL-2-transfected tumors. Thus, expression of IL-2 at the site of tumor growth may enhance tumor immunity not only by inducing the generation of tumor-reactive CTLs but also by allowing increased infiltration of activated T cells into the tumors. PMID:9537251

  9. Phenotypic and Functional Alterations of Dendritic Cells Induced by Human Herpesvirus 6 Infection

    OpenAIRE

    Kakimoto, Miki; Hasegawa, Atsuhiko; Fujita, Shigeru; Yasukawa, Masaki

    2002-01-01

    Human herpesvirus 6 (HHV-6) has a tropism for T lymphocytes and monocytes/macrophages, suggesting that HHV-6 infection affects the immunosurveillance system. In the present study, we investigated the HHV-6-induced phenotypic and functional alterations of dendritic cells (DCs), which are professional antigen-presenting cells. HHV-6 infection of monocyte-derived immature DCs appeared to induce the up-regulation of CD80, CD83, CD86, and HLA class I and class II molecules, suggesting that HHV-6 i...

  10. Alteration of B-cell subsets enhances neuroinvasion in mouse scrapie infection.

    Science.gov (United States)

    von Poser-Klein, Christine; Flechsig, Eckhard; Hoffmann, Tanja; Schwarz, Petra; Harms, Harry; Bujdoso, Raymond; Aguzzi, Adriano; Klein, Michael A

    2008-04-01

    Acquired forms of prion diseases or transmissible spongiform encephalopathies are believed to occur following peripheral exposure. Prions initially accumulate in the lymphoid system before spreading to the nervous system, but the underlying mechanisms for prion transfer between the two systems are still elusive. Here we show that ablation of the B-cell-specific transmembrane protein CD19, a coreceptor of the complement system, results in an acceleration of prion neuroinvasion. This appears to be due to an alteration of the follicular dendritic cell (FDC) network within the lymphoid tissue, thereby reducing the distance between FDCs and adjacent nerve fibers that mediate prion neuroinvasion. PMID:18199638

  11. Haemophilus haemolyticus Interaction with Host Cells Is Different to Nontypeable Haemophilus influenzae and Prevents NTHi Association with Epithelial Cells.

    Science.gov (United States)

    Pickering, Janessa L; Prosser, Amy; Corscadden, Karli J; de Gier, Camilla; Richmond, Peter C; Zhang, Guicheng; Thornton, Ruth B; Kirkham, Lea-Ann S

    2016-01-01

    Nontypeable Haemophilus influenzae (NTHi) is an opportunistic pathogen that resides in the upper respiratory tract and contributes to a significant burden of respiratory related diseases in children and adults. Haemophilus haemolyticus is a respiratory tract commensal that can be misidentified as NTHi due to high levels of genetic relatedness. There are reports of invasive disease from H. haemolyticus, which further blurs the species boundary with NTHi. To investigate differences in pathogenicity between these species, we optimized an in vitro epithelial cell model to compare the interaction of 10 H. haemolyticus strains with 4 NTHi and 4 H. influenzae-like haemophili. There was inter- and intra-species variability but overall, H. haemolyticus had reduced capacity to attach to and invade nasopharyngeal and bronchoalveolar epithelial cell lines (D562 and A549) within 3 h when compared with NTHi. H. haemolyticus was cytotoxic to both cell lines at 24 h, whereas NTHi was not. Nasopharyngeal epithelium challenged with some H. haemolyticus strains released high levels of inflammatory mediators IL-6 and IL-8, whereas NTHi did not elicit an inflammatory response despite higher levels of cell association and invasion. Furthermore, peripheral blood mononuclear cells stimulated with H. haemolyticus or NTHi released similar and high levels of IL-6, IL-8, IL-10, IL-1β, and TNFα when compared with unstimulated cells but only NTHi elicited an IFNγ response. Due to the relatedness of H. haemolyticus and NTHi, we hypothesized that H. haemolyticus may compete with NTHi for colonization of the respiratory tract. We observed that in vitro pre-treatment of epithelial cells with H. haemolyticus significantly reduced NTHi attachment, suggesting interference or competition between the two species is possible and warrants further investigation. In conclusion, H. haemolyticus interacts differently with host cells compared to NTHi, with different immunostimulatory and cytotoxic

  12. Characteristics of nobiletin-mediated alteration of gene expression in cultured cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Nemoto, Kiyomitsu, E-mail: nemoto@u-shizuoka-ken.ac.jp [Department of Molecular Toxicology, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526 (Japan); Ikeda, Ayaka; Yoshida, Chiaki; Kimura, Junko; Mori, Junki [Department of Molecular Toxicology, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526 (Japan); Fujiwara, Hironori [Department of Anti-Dementia Functional Food Development, Research Center of Supercritical Fluid Technology, Graduate School of Engineering, Tohoku University, 6-6-7 Aoba, Aramaki, Aoba-ku, Sendai 980-8579 (Japan); Yokosuka, Akihito; Mimaki, Yoshihiro [Department of Medicinal Pharmacognosy, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, 1432-1 Horinouchi, Hachioji 192-0392 (Japan); Ohizumi, Yasushi [Department of Molecular Toxicology, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526 (Japan); Department of Anti-Dementia Functional Food Development, Research Center of Supercritical Fluid Technology, Graduate School of Engineering, Tohoku University, 6-6-7 Aoba, Aramaki, Aoba-ku, Sendai 980-8579 (Japan); Laboratory of Kampo Medicines, Yokohama College of Pharmacy, 601 Matano-cho, Totsuka-ku, Yokohama 245-0066 (Japan); Degawa, Masakuni [Department of Molecular Toxicology, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526 (Japan)

    2013-02-15

    Highlights: ► Nobiletin-mediated alterations of gene expression were examined with DNA microarrays. ► Three organ-derived cell lines were treated with 100 μM nobiletin for 24 h. ► In all cell lines, 3 endoplasmic reticulum stress-responsive genes were up-regulated. ► Some cell cycle-regulating and oxidative stress-promoting genes were down-regulated. ► These alterations may contribute to nobiletin-mediated biological effects. -- Abstract: Nobiletin, a polymethoxylated flavonoid that is highly contained in the peels of citrus fruits, exerts a wide variety of beneficial effects, including anti-proliferative effects in cancer cells, repressive effects in hyperlipidemia and hyperglycemia, and ameliorative effects in dementia at in vitro and in vivo levels. In the present study, to further understand the mechanisms of these actions of nobiletin, the nobiletin-mediated alterations of gene expression in three organ-derived cell lines – 3Y1 rat fibroblasts, HuH-7 human hepatocarcinoma cells, and SK-N-SH human neuroblastoma cells – were first examined with DNA microarrays. In all three cell lines, treatments with nobiletin (100 μM) for 24 h resulted in more than 200% increases in the expression levels of five genes, including the endoplasmic reticulum stress-responsive genes Ddit3, Trib3, and Asns, and in less than 50% decreases in the expression levels of seven genes, including the cell cycle-regulating genes Ccna2, Ccne2, and E2f8 and the oxidative stress-promoting gene Txnip. It was also confirmed that in each nobiletin-treated cell line, the levels of the DDIT3 (DNA-damage-inducible transcript 3, also known as CHOP and GADD153) and ASNS (asparagine synthetase) proteins were increased, while the level of the TXNIP (thioredoxin-interacting protein, also known as VDUP1 and TBP-2) protein was decreased. All these findings suggest that nobiletin exerts a wide variety of biological effects, at least partly, through induction of endoplasmic reticulum stress and

  13. HIV Integration at Certain Sites in Host DNA Is Linked to the Expansion and Persistence of Infected Cells | Poster

    Science.gov (United States)

    Editor’s note: This article was originally published on the Center for Cancer Research website. When the Human Immunodeficiency Virus (HIV) infects a cell, the virus inserts a copy of its genetic material into the host cell’s DNA. The inserted genetic material, which is also called a provirus, is used to produce new viruses. Because the viral DNA can be inserted at many sites in the host cell DNA, the site of integration marks each infected cell. Patients infected with HIV are currently treated with combined antiretroviral therapy (cART), which prevents viral replication in the majority of treated patients. When cART is initiated, most HIV-infected cells die in one or two days, and more of the infected cells die over a period of weeks to months. However there are some long-lived infected cells that do not die, which prevents patients from being cured.

  14. Positional and expressive alteration of prohibitin during the induced differentiation of human hepatocarcinoma SMMC-7721 cells

    Institute of Scientific and Technical Information of China (English)

    Dong-Hui Xu; Jian Tang; Qi-Fu Li; Song-Lin Shi; Xiang-Feng Chen; Ying Liang

    2008-01-01

    AIM: To explore the existence and distribution of prohibitin (PHB) in nuclear matrix and its co-localization with products of some related genes during the differentiation of human hepatocarcinoma SMMC-7721cells.METHODS: The nuclear matrix of the SHHC-7721 cells cultured with or without 5 x 10-3 mmol/L hexamethylene bisacetamide (HMBA) was selectively extracted.Western blot was used to analyze the expression of PHB in nuclear matrix; imrnunofluorescence microscope observation was used to analyze the distribution of PHB in cell. LCSM was used to observe the co-localization of PHB with products of oncogenes and tumor suppressor genes.RESULTS: Western blot analysis showed that PHB existed in the composition of nuclear matrix proteins and was down-regulated by HMBA treatment.Immunofluorescence observation revealed that PHB existed in the nuclear matrix, and its distribution regions and expression levels were altered after HMBA treatment. Laser scanning confocal microscopy revealed the co-localization between PHB and the products of oncogenes or tumor repression genes including c-fos, c-myc, p53 and Rb and its alteration of distributive area in the cells treated by HMBA.CONCLUSION: These data confirm that PHB is a nuclear matrix protein, which is located in the nuclear matrix, and the distribution and expression of PHB and its relation with associated genes may play significant roles during the differentiation of SMHC-7721 cells.

  15. Alterations of p53 in tumorigenic human bronchial epithelial cells correlate with metastatic potential

    Science.gov (United States)

    Piao, C. Q.; Willey, J. C.; Hei, T. K.; Hall, E. J. (Principal Investigator)

    1999-01-01

    The cellular and molecular mechanisms of radiation-induced lung cancer are not known. In the present study, alterations of p53 in tumorigenic human papillomavirus-immortalized human bronchial epithelial (BEP2D) cells induced by a single low dose of either alpha-particles or 1 GeV/nucleon (56)Fe were analyzed by PCR-single-stranded conformation polymorphism (SSCP) coupled with sequencing analysis and immunoprecipitation assay. A total of nine primary and four secondary tumor cell lines, three of which were metastatic, together with the parental BEP2D and primary human bronchial epithelial (NHBE) cells were studied. The immunoprecipitation assay showed overexpression of mutant p53 proteins in all the tumor lines but not in NHBE and BEP2D cells. PCR-SSCP and sequencing analysis found band shifts and gene mutations in all four of the secondary tumors. A G-->T transversion in codon 139 in exon 5 that replaced Lys with Asn was detected in two tumor lines. One mutation each, involving a G-->T transversion in codon 215 in exon 6 (Ser-->lle) and a G-->A transition in codon 373 in exon 8 (Arg-->His), was identified in the remaining two secondary tumors. These results suggest that p53 alterations correlate with tumorigenesis in the BEP2D cell model and that mutations in the p53 gene may be indicative of metastatic potential.

  16. Mass spectrometric analysis of host cell proteins interacting with dengue virus nonstructural protein 1 in dengue virus-infected HepG2 cells.

    Science.gov (United States)

    Dechtawewat, Thanyaporn; Paemanee, Atchara; Roytrakul, Sittiruk; Songprakhon, Pucharee; Limjindaporn, Thawornchai; Yenchitsomanus, Pa-Thai; Saitornuang, Sawanan; Puttikhunt, Chunya; Kasinrerk, Watchara; Malasit, Prida; Noisakran, Sansanee

    2016-09-01

    Dengue virus (DENV) infection is a leading cause of the mosquito-borne infectious diseases that affect humans worldwide. Virus-host interactions appear to play significant roles in DENV replication and the pathogenesis of DENV infection. Nonstructural protein 1 (NS1) of DENV is likely involved in these processes; however, its associations with host cell proteins in DENV infection remain unclear. In this study, we used a combination of techniques (immunoprecipitation, in-solution trypsin digestion, and LC-MS/MS) to identify the host cell proteins that interact with cell-associated NS1 in an in vitro model of DENV infection in the human hepatocyte HepG2 cell line. Thirty-six novel host cell proteins were identified as potential DENV NS1-interacting partners. A large number of these proteins had characteristic binding or catalytic activities, and were involved in cellular metabolism. Coimmunoprecipitation and colocalization assays confirmed the interactions of DENV NS1 and human NIMA-related kinase 2 (NEK2), thousand and one amino acid protein kinase 1 (TAO1), and component of oligomeric Golgi complex 1 (COG1) proteins in virus-infected cells. This study reports a novel set of DENV NS1-interacting host cell proteins in the HepG2 cell line and proposes possible roles for human NEK2, TAO1, and COG1 in DENV infection. PMID:27108190

  17. Erythropoietin withdrawal alters interactions between young red blood cells, splenic endothelial cells, and macrophages: an in vitro model of neocytolysis

    Science.gov (United States)

    Trial, J.; Rice, L.; Alfrey, C. P.

    2001-01-01

    BACKGROUND: We have described the rapid destruction of young red blood cells (neocytolysis) in astronauts adapting to microgravity, in polycythemic high altitude dwellers who descend to sea level, and in patients with kidney disorders. This destruction results from a decrease in erythropoietin (EPO) production. We hypothesized that such EPO withdrawal could trigger physiological changes in cells other than red cell precursors and possibly lead to the uptake and destruction of young red cells by altering endothelial cell-macrophage interactions, most likely occurring in the spleen. METHODS: We identified EPO receptors on human splenic endothelial cells (HSEC) and investigated the responses of these cells to EPO withdrawal. RESULTS: A monolayer of HSEC, unlike human endothelial cells from aorta, glomerulus, or umbilical vein, demonstrated an increase in permeability upon EPO withdrawal that was accompanied by unique morphological changes. When HSEC were cultured with monocyte-derived macrophages (but not when either cell type was cultured alone), EPO withdrawal induced an increased ingestion of young red cells by macrophages when compared with the constant presence or absence of EPO. CONCLUSIONS: HSEC may represent a unique cell type that is able to respond to EPO withdrawal by increasing permeability and interacting with phagocytic macrophages, which leads to neocytolysis.

  18. Phospholipase D promotes Arcanobacterium haemolyticum adhesion via lipid raft remodeling and host cell death following bacterial invasion

    Directory of Open Access Journals (Sweden)

    Carlson Petteri

    2010-10-01

    Full Text Available Abstract Background Arcanobacterium haemolyticum is an emerging bacterial pathogen, causing pharyngitis and more invasive infections. This organism expresses an unusual phospholipase D (PLD, which we propose promotes bacterial pathogenesis through its action on host cell membranes. The pld gene is found on a genomic region of reduced %G + C, suggesting recent horizontal acquisition. Results Recombinant PLD rearranged HeLa cell lipid rafts in a dose-dependent manner and this was inhibited by cholesterol sequestration. PLD also promoted host cell adhesion, as a pld mutant had a 60.3% reduction in its ability to adhere to HeLa cells as compared to the wild type. Conversely, the pld mutant appeared to invade HeLa cells approximately two-fold more efficiently as the wild type. This finding was attributable to a significant loss of host cell viability following secretion of PLD from intracellular bacteria. As determined by viability assay, only 15.6% and 82.3% of HeLa cells remained viable following invasion by the wild type or pld mutant, respectively, as compared to untreated HeLa cells. Transmission electron microscopy of HeLa cells inoculated with A. haemolyticum strains revealed that the pld mutant was contained within intracellular vacuoles, as compared to the wild type, which escaped the vacuole. Wild type-infected HeLa cells also displayed the hallmarks of necrosis. Similarly inoculated HeLa cells displayed no signs of apoptosis, as measured by induction of caspase 3/7, 8 or 9 activities. Conclusions These data indicate that PLD enhances bacterial adhesion and promotes host cell necrosis following invasion, and therefore, may be important in the disease pathogenesis of A. haemolyticum infections.

  19. Allogeneic Th1 Cells Home to Host Bone Marrow and Spleen and Mediate IFNγ-Dependent Aplasia

    OpenAIRE

    Chewning, Joseph H.; Zhang, Weiwei; Randolph, David A.; Swindle, C. Scott; Schoeb, Trenton R.; Weaver, Casey T.

    2013-01-01

    Bone marrow graft failure and poor graft function are frequent complications following hematopoietic stem cell transplantation and result in significant morbidity and mortality. Both conditions are associated with graft versus host disease (GVHD), although the mechanism remains undefined. Here we show in two distinct murine models of GVHD (complete MHC- and class II-disparate) that mimic human peripheral blood stem cell transplantation that Th1 CD4+ cells induce bone marrow failure in allogen...

  20. Anaplerotic Metabolism of Alloreactive T Cells Provides a Metabolic Approach To Treat Graft-Versus-Host Disease

    OpenAIRE

    Glick, Gary D.; Rossignol, Rodrigue; Lyssiotis, Costas A.; Wahl, Daniel; Lesch, Charles; Sanchez, Brian; Liu, Xikui; Hao, Ling-Yang; Taylor, Clarke; Hurd, Alexander; Ferrara, James L. M.; Tkachev, Victor; Byersdorfer, Craig A.; Boros, Laszlo; Opipari, Anthony W.

    2014-01-01

    T-cell activation requires increased ATP and biosynthesis to support proliferation and effector function. Most models of T-cell activation are based on in vitro culture systems and posit that aerobic glycolysis is employed to meet increased energetic and biosynthetic demands. By contrast, T cells activated in vivo by alloantigens in graft-versus-host disease (GVHD) increase mitochondrial oxygen consumption, fatty acid uptake, and oxidation, with small increases of glucose uptake and aerobic g...

  1. Tracing Conidial Fate and Measuring Host Cell Antifungal Activity Using a Reporter of Microbial Viability in the Lung

    OpenAIRE

    Jhingran, Anupam; Mar, Katrina B.; Kumasaka, Debra K.; Sue E Knoblaugh; Ngo, Lisa Y.; Segal, Brahm H; Iwakura, Yoichiro; Lowell, Clifford A.; Hamerman, Jessica A.; Lin, Xin; Tobias M Hohl

    2012-01-01

    Fluorescence can be harnessed to monitor microbial fate and to investigate functional outcomes of individual microbial cell-host cell encounters at portals of entry in native tissue environments. We illustrate this concept by introducing fluorescent Aspergillus reporter (FLARE) conidia that simultaneously report phagocytic uptake and fungal viability during cellular interactions with the murine respiratory innate immune system. Our studies using FLARE conidia reveal stepwise and cell-type-spe...

  2. Altered cell wall disassembly during ripening of Cnr tomato fruit: implications for cell adhesion and fruit softening

    DEFF Research Database (Denmark)

    Orfila, C.; Huisman, M.M.H.; Willats, William George Tycho;

    2002-01-01

    The Cnr (Colourless non-ripening) tomato (Lycopersicon esculentum Mill.) mutant has an aberrant fruit-ripening phenotype in which fruit do not soften and have reduced cell adhesion between pericarp cells. Cell walls from Cnr fruit were analysed in order to assess the possible contribution of pectic...... polysaccharides to the non-softening and altered cell adhesion phenotype. Cell wall material (CWM) and solubilised fractions of mature green and red ripe fruit were analysed by chemical, enzymatic and immunochemical techniques. No major differences in CWM sugar composition were detected although differences were...... found in the solubility and composition of the pectic polysaccharides extracted from the CWM at both stages of development. In comparison with the wild type, the ripening-associated solubilisation of homogalacturonan-rich pectic polysaccharides was reduced in Cnr. The proportion of carbohydrate...

  3. Bst1 is required for Candida albicans infecting host via facilitating cell wall anchorage of Glycosylphosphatidyl inositol anchored proteins

    Science.gov (United States)

    Liu, Wei; Zou, Zui; Huang, Xin; Shen, Hui; He, Li Juan; Chen, Si Min; Li, Li Ping; Yan, Lan; Zhang, Shi Qun; Zhang, Jun Dong; Xu, Zheng; Xu, Guo Tong; An, Mao Mao; Jiang, Yuan Ying

    2016-01-01

    Glycosylphosphatidyl inositol anchored proteins (GPI-APs) on fungal cell wall are essential for invasive infections. While the function of inositol deacylation of GPI-APs in mammalian cells has been previously characterized the impact of inositol deacylation in fungi and implications to host infection remains largely unexplored. Herein we describe our identification of BST1, an inositol deacylase of GPI-Aps in Candida albicans, was critical for GPI-APs cell wall attachment and host infection. BST1-deficient C. albicans (bst1Δ/Δ) was associated with severely impaired cell wall anchorage of GPI-APs and subsequen unmasked β-(1,3)-glucan. Consistent with the aberrant cell wall structures, bst1Δ/Δ strain did not display an invasive ability and could be recognized more efficiently by host immune systems. Moreover, BST1 null mutants or those expressing Bst1 variants did not display inositol deacylation activity and exhibited severely attenuated virulence and reduced organic colonization in a murine systemic candidiasis model. Thus, Bst1 can facilitate cell wall anchorage of GPI-APs in C. albicans by inositol deacylation, and is critical for host invasion and immune escape. PMID:27708385

  4. Systemic Sclerosis Patients Present Alterations in the Expression of Molecules Involved in B-Cell Regulation

    Science.gov (United States)

    Soto, Lilian; Ferrier, Ashley; Aravena, Octavio; Fonseca, Elianet; Berendsen, Jorge; Biere, Andrea; Bueno, Daniel; Ramos, Verónica; Aguillón, Juan Carlos; Catalán, Diego

    2015-01-01

    The activation threshold of B cells is tightly regulated by an array of inhibitory and activator receptors in such a way that disturbances in their expression can lead to the appearance of autoimmunity. The aim of this study was to evaluate the expression of activating and inhibitory molecules involved in the modulation of B cell functions in transitional, naive, and memory B-cell subpopulations from systemic sclerosis patients. To achieve this, blood samples were drawn from 31 systemic sclerosis patients and 53 healthy individuals. Surface expression of CD86, MHC II, CD19, CD21, CD40, CD22, Siglec 10, CD35, and FcγRIIB was determined by flow cytometry. IL-10 production was evaluated by intracellular flow cytometry from isolated B cells. Soluble IL-6 and IL-10 levels were measured by ELISA from supernatants of stimulated B cells. Systemic sclerosis patients exhibit an increased frequency of transitional and naive B cells related to memory B cells compared with healthy controls. Transitional and naive B cells from patients express higher levels of CD86 and FcγRIIB than healthy donors. Also, B cells from patients show high expression of CD19 and CD40, whereas memory cells from systemic sclerosis patients show reduced expression of CD35. CD19 and CD35 expression levels associate with different autoantibody profiles. IL-10+ B cells and secreted levels of IL-10 were markedly reduced in patients. In conclusion, systemic sclerosis patients show alterations in the expression of molecules involved in B-cell regulation. These abnormalities may be determinant in the B-cell hyperactivation observed in systemic sclerosis. PMID:26483788

  5. Microgravity-induced alterations in signal transduction in cells of the immune system

    Science.gov (United States)

    Paulsen, Katrin; Thiel, Cora; Timm, Johanna; Schmidt, Peter M.; Huber, Kathrin; Tauber, Svantje; Hemmersbach, Ruth; Seibt, Dieter; Kroll, Hartmut; Grote, Karl-Heinrich; Zipp, Frauke; Schneider-Stock, Regine; Cogoli, Augusto; Hilliger, Andre; Engelmann, Frank; Ullrich, Oliver

    2010-11-01

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