cholesteryl-glucoside and Helicobacter-Infections

Steryl glycosides are derivatives of sterols where the 3β-hydroxy group is glycosylated. Some of them are further converted to steryl O-acyl glycosides. Steryl glycosides and their derivatives are widely distributed in plants, algae, and fungi, but are relatively rarely distributed in bacteria and animals. Accumulating evidence suggests that glycosylation of sterols not only modifies physicochemical properties of cell membranes but also alters immunogenicity of the cells. Helicobacter pylori, that colonizes the stomach and causes gastric diseases, is auxotrophic for cholesterol, so that it extracts this lipid from plasma membranes of epithelial cells of the host stomach. Since incorporation of cholesterol promotes immune responses of the host, Helicobacter pylori converts cholesterol to cholesteryl glucoside (ChG) and then to cholesteryl 6'-O-acyl glucoside (ChAcG) to evade the immune surveillance. We have found that ChAcG thus produced is specifically recognized by invariant Vα14-Jα18 TCR(+) (Vα14) NKT cells in a CD1-dependent manner. We have also found that activation of Vα14 NKT cells by administration of ChAcG retains homeostasis of immunity upon exposure to allergens and reduces the incidence of allergy. In this article, overview of immunological functions of steryl glycosides with an emphasis on the immunoregulatory functions of ChAcG, is demonstrated.

Topics: Animals; Antigens, CD1; Cholesterol; Cytokines; Helicobacter Infections; Helicobacter pylori; Humans; Natural Killer T-Cells; Th1 Cells

During autophagy defense against invading microbes, certain lipid types are indispensable for generating specialized membrane-bound organelles. The lipid composition of autophagosomes remains obscure, as does the issue of how specific lipids and lipid-associated enzymes participate in autophagosome formation and maturation. Helicobacter pylori is auxotrophic for cholesterol and converts cholesterol to cholesteryl glucoside derivatives, including cholesteryl 6'-O-acyl-α-D-glucoside (CAG). We investigated how CAG and its biosynthetic acyltransferase assist H. pylori to escape host-cell autophagy.. We applied a metabolite-tagging method to obtain fluorophore-containing cholesteryl glucosides that were utilized to understand their intracellular locations. H. pylori 26695 and a cholesteryl glucosyltransferase (CGT)-deletion mutant (ΔCGT) were used as the standard strain and the negative control that contains no cholesterol-derived metabolites, respectively. Bacterial internalization and several autophagy-related assays were conducted to unravel the possible mechanism that H. pylori develops to hijack the host-cell autophagy response. Subcellular fractions of H. pylori-infected AGS cells were obtained and measured for the acyltransferase activity.. The imaging studies of fluorophore-labeled cholesteryl glucosides pinpointed their intracellular localization in AGS cells. The result indicated that CAG enhances the internalization of H. pylori in AGS cells. Particularly, CAG, instead of CG and CPG, is able to augment the autophagy response induced by H. pylori. How CAG participates in the autophagy process is multifaceted. CAG was found to intervene in the degradation of autophagosomes and reduce lysosomal biogenesis, supporting the idea that intracellular H. pylori is harbored by autophago-lysosomes in favor of the bacterial survival. Furthermore, we performed the enzyme activity assay of subcellular fractions of H. pylori-infected AGS cells. The analysis showed that the acyltransferase is mainly distributed in autophago-lysosomal compartments.. Our results support the idea that the acyltransferase is mainly distributed in the subcellular compartment consisting of autophagosomes, late endosomes, and lysosomes, in which the acidic environment is beneficial for the maximal acyltransferase activity. The resulting elevated level of CAG can facilitate bacterial internalization, interfere with the autophagy flux, and causes reduced lysosomal biogenesis.

Topics: Acyltransferases; Animals; Cholesterol; Helicobacter Infections; Helicobacter pylori; Lysosomes; Male; Mice; Mice, Inbred C57BL; Specific Pathogen-Free Organisms

Helicobacter pylori colonizes the human stomach and contributes to chronic inflammation of the gastric mucosa. H. pylori persistence occurs because of insufficient eradication by phagocytic cells. A key factor of H. pylori, cholesterol-α-glucosyltransferase encoded by capJ that extracts host cholesterol and converts it to cholesteryl glucosides, is important to evade host immunity. Here, we examined whether phagocytic trafficking in macrophages was perturbed by capJ-carrying H. pylori.. J774A.1 cells were infected with H. pylori at a multiplicity of infection of 50. Live-cell imaging and confocal microscopic analysis were applied to monitor the phagocytic trafficking events. The viability of H. pylori inside macrophages was determined by using gentamicin colony-forming unit assay. The phagocytic routes were characterized by using trafficking-intervention compounds.. Wild type (WT) H. pylori exhibited more delayed entry into macrophages and also arrested phagosome maturation more than did capJ knockout mutant. Pretreatment of genistein and LY294002 prior to H. pylori infection reduced the internalization of WT but not capJ-knockout H. pylori in macrophages.. Cholesterol glucosylation by H. pylori interferes with phagosome trafficking via a lipid-raft and PI3K-dependent manner, which retards engulfment of bacteria for prolonged intracellular survival of H. pylori.

Topics: Animals; Cell Line; Cholesterol; Glucosyltransferases; Helicobacter Infections; Helicobacter pylori; Immune Evasion; Macrophages; Membrane Microdomains; Mice; Microscopy, Confocal; Phagocytosis; Phagosomes; Phosphatidylinositol 3-Kinases

Steryl glycosides produced by bacteria play important biological roles in the evasion and modulation of host immunity. Step-economical syntheses of three cholesteryl-6-O-phosphatidyl-α-D-glucopyranosides (αCPG) unique to Helicobacter pylori have been achieved. The approach relies upon regioselective deprotection of per-O-trimethylsilyl-α-D-cholesterylglucoside at C6 followed by phosphoramidite coupling. Global TMS ether deprotection in the presence of oxygen and subsequent deprotection of the cyano ethyl phosphoester afforded the target compounds in 16-21 % overall yield starting from D-glucose. The structures of these natural products were determined using a combination of 2D NMR methods and mass spectrometry. These robust synthesis and characterization protocols provide analogues to facilitate glycolipidomic profiling and biological studies.

Topics: Chemistry Techniques, Synthetic; Cholesterol; Helicobacter Infections; Helicobacter pylori; Humans; Immunologic Factors; Phosphorylation

Helicobacter pylori infection is an aetiological cause of gastric disorders worldwide. H. pylori has been shown to assimilate and convert host cholesterol into cholesteryl glucosides (CGs) by cholesterol-α-glucosyltransferase encoded by capJ. Here, we show that CapJ-deficient (ΔcapJ) H. pylori resulted in greatly reduced type IV secretion system (TFSS)-associated activities, including the hummingbird phenotype of AGS cells, IL-8 production, CagA translocation/phosphorylation and CagA-mediated signalling events. Complementation of the ΔcapJ mutation with wild type cagJ or by adding CGs-containing lysates or exogenous fluorophore-tagged CGs reversed the mutant phenotypes. We also show that the wild-type but not ΔcapJ H. pylori recruited raft-associated components to sites of bacterial attachment. Fluorescence recovery after photobleaching (FRAP) analysis of AGS cells treated with fluorescence-tagged cholesterol/CGs revealed that there was a higher proportion of CGs associated with immobile fractions. CGs-associated membranes were also more resistant to a cold detergent extraction. Thus, we propose that CGs synthesized by H. pylori around host-pathogen contact sites partition in detergent-resistant membranes (DRMs), alters lateral-phase segregation in membrane and reorganizes membrane architecture. These processes together promote the formation of a functional TFSS and H. pylori infection.

Topics: Antigens, Bacterial; Bacterial Proteins; Bacterial Secretion Systems; Cell Line, Tumor; Cell Membrane; Cholesterol; Helicobacter Infections; Helicobacter pylori; Humans

Understanding sporadic cases of age-dependent neurodegenerative diseases such as parkinsonism requires the evaluation of potential environmental factors. Amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS-PDC), a neurological disorder in which features of parkinsonism are present and for which no consistent genetic explanation has been found, has been linked to the consumption of cycad (Cycas micronesica). Similarly, epidemiological evidence suggests an association between parkinsonism and gastric ulcer caused by Helicobacter pylori infection. While common immunological and inflammatory changes have been proposed to account for the link between parkinsonism and H. pylori infection, we propose an alternate explanation based on our work on the "cycad theory" of ALS-PDC. Recent experiments in our laboratory have identified several sterol glucosides in cycad that have neurotoxic properties in vitro and that appear to be linked to the development of neurodegenerative disease in vivo. Specifically, mice fed cycad display behavioural symptoms of parkinsonism such as reduced gait length, as well as neuropathological signs such as a loss of striatal dopaminergic (DAergic) terminals and an upregulation of the dopamine D2 receptor. These cycad-derived sterol glucosides are structurally similar to cholesterol glucosides that account for a significant part pf the lipid profile of H. pylori. We hypothesize that cholesterol glucosides arising from H. pylori infection may act as neurotoxins, promoting the degeneration of the DAergic neurons affected in parkinsonism, in a similar reaction to that which is thought to link cycad consumption and ALS-PDC. This hypothesis will be tested in future studies that will include exposing mice to purified sterol or cholestorol glucosides derived from cycad and comparing these mice behaviourally and neuropathologically to ones chronically infected with H. pylori.

Topics: Amyotrophic Lateral Sclerosis; Cholesterol; Cycas; Helicobacter Infections; Helicobacter pylori; Humans; Models, Biological; Parkinson Disease; Plants, Toxic