Maria Kaparakis-Liaskos

Publications of Maria Kaparakis-Liaskos

• The use of AlbuMAX II(®) as a blood or serum alternative for the culture of Helicobacter pylori.

  Authors: Melanie L Hutton, Maria Kaparakis-Liaskos, Richard L Ferrero

  Helicobacter. 02/2012; 17(1):68-76.

  Growth of Helicobacter pyloriin vitro depends on supplementation of the medium with blood or serum. However, these supplements often require frozen storage and can show batch-to-batch variation,Growth of Helicobacter pyloriin vitro depends on supplementation of the medium with blood or serum. However, these supplements often require frozen storage and can show batch-to-batch variation, resulting in differences in bacterial growth. In this study, we introduce the use of a commercially available, lipid-rich supplement called AlbuMAX II(®) (Gibco BRL, Grand Island, NY, USA) for use as a serum/blood replacement for H. pylori culture. The growth of H. pylori on solid and liquid media was examined by comparing growth after supplementation with horse blood, fetal calf serum, β-cyclodextrin or AlbuMAX II(®) (Gibco BRL). Human gastric adenocarcinoma (AGS) cellular responses to H. pylori were measured by NF-κB luciferase assays and IL-8 ELISA. We show that the growth of H. pylori on both solid and liquid media containing AlbuMAX II(®) (Gibco BRL) were comparable to levels obtained on blood agar or liquid media supplemented with serum. Growth was consistently higher in media supplemented with AlbuMAX II(®) (Gibco BRL) than media containing β-cyclodextrin. Furthermore, bacteria grown in AlbuMAX II(®) (Gibco BRL) induced proinflammatory responses in AGS cells. AlbuMAX II(®) (Gibco BRL) can be used as a serum/blood replacement for the cultivation of H. pylori in solid and liquid media. This medium could be useful for an improved understanding of H. pylori metabolism or for antigen production. Furthermore, AlbuMAX II(®) (Gibco BRL) may be suitable for use in remote locations, particularly in areas where frozen storage of serum may be a problem.

• Helicobacter pylori exploits cholesterol-rich microdomains for induction of NF-kappaB-dependent responses and peptidoglycan delivery in epithelial cells.

  Authors: Melanie L Hutton, Maria Kaparakis-Liaskos, Lorinda Turner, Ana Cardona, Terry Kwok, Richard L Ferrero

  Infection and immunity. 11/2010; 78(11):4523-31.

  Infection with Helicobacter pylori cag pathogenicity island (cagPAI)-positive strains is associated with more destructive tissue damage and an increased risk of severe disease. The cagPAI encodes aInfection with Helicobacter pylori cag pathogenicity island (cagPAI)-positive strains is associated with more destructive tissue damage and an increased risk of severe disease. The cagPAI encodes a type IV secretion system (TFSS) that delivers the bacterial effector molecules CagA and peptidoglycan into the host cell cytoplasm, thereby inducing responses in host cells. It was previously shown that interactions between CagL, present on the TFSS pilus, and host α(5)β(1) integrin molecules were critical for CagA translocation and the induction of cytoskeletal rearrangements in epithelial cells. As the α(5)β(1) integrin is found in cholesterol-rich microdomains (known as lipid rafts), we hypothesized that these domains may also be involved in the induction of proinflammatory responses mediated by NOD1 recognition of H. pylori peptidoglycan. Indeed, not only did methyl-β-cyclodextrin depletion of cholesterol from cultured epithelial cells have a significant effect on the levels of NF-κB and interleukin-8 (IL-8) responses induced by H. pylori bacteria with an intact TFSS (P < 0.05), but it also interfered with TFSS-mediated peptidoglycan delivery to cells. Both of these effects could be restored by cholesterol replenishment of the cells. Furthermore, we demonstrated for the first time the involvement of α(5)β(1) integrin in the induction of proinflammatory responses by H. pylori. Taking the results together, we propose that α(5)β(1) integrin, which is associated with cholesterol-rich microdomains at the host cell surface, is required for NOD1 recognition of peptidoglycan and subsequent induction of NF-κB-dependent responses to H. pylori. These data implicate cholesterol-rich microdomains as a novel platform for TFSS-dependent delivery of bacterial products to cytosolic pathogen recognition molecules.

• Genetic modulation of TLR8 response following bacterial phagocytosis.

  Authors: Michael P Gantier, Aaron T Irving, Maria Kaparakis-Liaskos, Dakang Xu, Vanessa A Evans, Paul U Cameron, James A Bourne, Richard L Ferrero, Matthias John, Mark A Behlke, Bryan R G Williams

  Human mutation. 09/2010; 31(9):1069-79.

  Human Toll-like receptors (TLRs) TLR7, TLR8, and TLR9 are important immune sensors of foreign nucleic acids encountered by phagocytes. Although there is growing evidence implicating TLR7 and TLR9 inHuman Toll-like receptors (TLRs) TLR7, TLR8, and TLR9 are important immune sensors of foreign nucleic acids encountered by phagocytes. Although there is growing evidence implicating TLR7 and TLR9 in the detection of intracellular pathogenic bacteria, characterization of such a role for TLR8 is currently lacking. A recent genetic study has correlated the presence of a TLR8 single nucleotide polymorphism (SNP) (rs3764880:A>G; p.Met1Val) with the development of active tuberculosis, suggesting a role for TLR8 in the detection of phagosomal bacteria. Here we provide the first direct evidence that TLR8 sensing is activated in human monocytic cells following Helicobacter pylori phagocytosis. In addition, we show that rs3764880 fine tunes translation of the two TLR8 main isoforms, without affecting protein function. Although we show that TLR8 variant 2 (TLR8v2) is the prevalent form of TLR8 contributing to TLR8 function, we also uncover a role for the TLR8 long isoform (TLR8v1) in the positive regulation of TLR8 function in CD16(+)CD14(+) differentiated monocytes. Thus, TLR8 sensing can be activated following bacterial phagocytosis, and rs3764880 may play a role in the modulation of TLR8-dependent microbicidal response of infected macrophages.

• Vitamin B6 is required for full motility and virulence in Helicobacter pylori.

  Authors: Alexandra Grubman, Alexandra Phillips, Marie Thibonnier, Maria Kaparakis-Liaskos, Chad Johnson, Jean-Michel Thiberge, Fiona J Radcliff, Chantal Ecobichon, Agnès Labigne, Hilde de Reuse, George L Mendz, Richard L Ferrero

  mBio. 01/2010; 1(3).

  Despite recent advances in our understanding of how Helicobacter pylori causes disease, the factors that allow this pathogen to persist in the stomach have not yet been fully characterized. ToDespite recent advances in our understanding of how Helicobacter pylori causes disease, the factors that allow this pathogen to persist in the stomach have not yet been fully characterized. To identify new virulence factors in H. pylori, we generated low-infectivity variants of a mouse-colonizing H. pylori strain using the classical technique of in vitro attenuation. The resulting variants and their highly infectious progenitor bacteria were then analyzed by global gene expression profiling. The gene expression levels of five open reading frames (ORFs) were significantly reduced in low-infectivity variants, with the most significant changes observed for ORFs HP1583 and HP1582. These ORFs were annotated as encoding homologs of the Escherichia coli vitamin B(6) biosynthesis enzymes PdxA and PdxJ. Functional complementation studies with E. coli confirmed H. pylori PdxA and PdxJ to be bona fide homologs of vitamin B(6) biosynthesis enzymes. Importantly, H. pylori PdxA was required for optimal growth in vitro and was shown to be essential for chronic colonization in mice. In addition to having a well-known metabolic role, vitamin B(6) is necessary for the synthesis of glycosylated flagella and for flagellum-based motility in H. pylori. Thus, for the first time, we identify vitamin B(6) biosynthesis enzymes as novel virulence factors in bacteria. Interestingly, pdxA and pdxJ orthologs are present in a number of human pathogens, but not in mammalian cells. We therefore propose that PdxA/J enzymes may represent ideal candidates for therapeutic targets against bacterial pathogens.

• Helicobacter pylori-induced histone modification, associated gene expression in gastric epithelial cells, and its implication in pathogenesis.

  Authors: Song-Ze Ding, Wolfgang Fischer, Maria Kaparakis-Liaskos, George Liechti, D Scott Merrell, Patrick A Grant, Richard L Ferrero, Sheila E Crowe, Rainer Haas, Masanori Hatakeyama, Joanna B Goldberg

  PloS one. 01/2010; 5(4):e9875.

  Histone modifications are critical in regulating gene expression, cell cycle, cell proliferation, and development. Relatively few studies have investigated whether Helicobacter pylori, the majorHistone modifications are critical in regulating gene expression, cell cycle, cell proliferation, and development. Relatively few studies have investigated whether Helicobacter pylori, the major cause of human gastric diseases, affects histone modification. We therefore investigated the effects of H. pylori infection on histone modifications in a global and promoter-specific manner in gastric epithelial cells. Infection of gastric epithelial cells by wild-type H. pylori induced time- and dose-dependent dephosphorylation of histone H3 at serine 10 (H3 Ser10) and decreased acetylation of H3 lysine 23, but had no effects on seven other specific modifications. Different cag pathogenicity island (PAI)-containing-clinical isolates showed similar abilities to induce H3 Ser10 dephosphorylation. Mutation of cagA, vacA, nonphosphorylateable CagA mutant cagA(EPISA), or disruption of the flagella showed no effects, while deletion of the entire cagPAI restored the H3 Ser10 phosphorylation to control levels. Analysis of 27 cagPAI mutants indicated that the genes that caused H3 Ser10 dephosphorylation were similar to those that were previously found to induce interleukin-8, irrespective of CagA translocation. This effect was independent of ERK or p38 pathways and type I interferon signaling. Additionally, c-Jun and hsp70 gene expression was associated with this histone modification. These results demonstrate that H. pylori alters histone modification and host response via a cagA-, vacA-independent, but cagPAI-dependent mechanisms, which contribute to its persistent infection and pathogenesis.