Showing papers on "Phagosome published in 1985"
TL;DR: It is reported here that modified live Toxoplasma-containing vacuoles fail to acidify in normal macrophages, as indicated by the sensitive pH probe fluorescein.
Abstract: Toxoplasma gondii belongs to a group of highly virulent intracellular parasites that reside in host cell vacuoles which resist typical phagosome-lysosome fusion. Live Toxoplasma replicate prodigiously within modified phagocytic vacuoles formed during invagination of the host plasma membrane. In contrast, heat-killed Toxoplasma or specific antibody (heat-inactivated)-coated live Toxoplasma-containing vacuoles readily undergo lysosome fusion and digestion in normal macrophages. Of newly recognized significance to Toxoplasma survival is the microbicidal effect of phagosome acidification, which reportedly can occur independently of fusion with other acidic vesicles. We report here that modified live Toxoplasma-containing vacuoles fail to acidify in normal macrophages, as indicated by the sensitive pH probe fluorescein. In contrast, when live Toxoplasma are coated with specific antibody (heat-inactivated), they trigger phagosome acidification when entering normal macrophages. A similar acidification is observed when normal phagocytes ingest dead Toxoplasma. Extracellular Toxoplasma are highly susceptible to acidic pH conditions, indicating that the acidification block in the modified vacuoles may be important for intracellular survival.
284 citations
Journal Article•
TL;DR: Water extracts of virulent Brucella abortus were able to inhibit phagosome-lysosome fusion in unelicited murine peritoneal macrophages following ingestion of yeast.
Abstract: Water extracts of virulent Brucella abortus were able to inhibit phagosome-lysosome fusion in unelicited murine peritoneal macrophages following ingestion of yeast. Extracts from an avirulent strain were unable to produce a similar effect. Lipopolysaccharide from B abortus did not appear to be involved with the ability of the extracts to inhibit fusion.
96 citations
TL;DR: In vitro testing showed that ferritin dissociation requires an acid pH, indicating that phagosomes acidify prior to lysosome fusion, and Phagocytized Glugea hertwigi spores underwent acidification during this time period.
Abstract: This study examines the relationship between phagosome acidification and phagosome-lysosome fusion events using phagocytized Glugea hertwigi spores. The incidence of lysosome fusion with Glugea spores in phagosomes of mouse peritoneal macrophages and of Tetrahymena was monitored using colloidal gold and acridine orange as labels for secondary lysosomes. Over 80% of the Glugea phagosomes remained segregated from the labeled compartments in macrophages after 60 min; this inhibition of fusion was still evident after 4 h. In Tetrahymena, Glugea spores also showed a high capacity to block fusion with secondary lysosomes (67%); however, spores coated with cationized ferritin showed an 80% fusion rate with labeled acidic compartments (i.e. lysosomes) after 60 min with both Tetrahymena and macrophages. The pH of phagosome compartments was monitored by measuring the emissions of fluorescein isothiocyanate (FITC)-labeled Glugea ingested by Tetrahymena. Tetrahymena phagosomes with FITC-Glugea did not acidify within the first hour after phagocytosis; however, phagosomes with cationized ferritin-labeled Glugea underwent acidification during this time period. This acidification took place although the capability of the host cells' lysosomes to fuse was blocked by pretreatment with poly-D-glutamic acid. The cationized ferritin bound to Glugea spores was uncoupled from the spore wall prior to fusion with colloidal gold-labeled compartments. In vitro testing showed that ferritin dissociation requires an acid pH, indicating that phagosomes acidify prior to lysosome fusion.
61 citations
TL;DR: The main function of the macrophages, which is to ingest and degrade any foreign molecules or particles penetrating the organism, appears in the development of the different structures implicated in endocytic activity, and endocytosis is accompanied by rapid and intense exchanges between the different membrane compartments of the cell.
Abstract: The main function of the macrophages, which is to ingest and degrade any foreign molecules or particles penetrating the organism, appears in the development of the different structures implicated in endocytic activity. The macrophage's high endocytic property first appears in its irregular shape and the large number of extensions of the cell membrane, allowing the rapid capture of extra-cellular material. Adhesion between macrophage cell surface and molecules or particles is greatly enhanced by the presence of varied kinds of receptors: lectin-like receptors which bind specific sugars or highly specific receptors such as Fc and C3b receptors, which increase phagocytosis of opsonized microbes. The microbicidal properties reside in part in the production of superoxide anions which result from the activity of a NAD(P)H oxidase. This enzyme is located in the plasma membrane. Its activity could be demonstrated with a cytochemical method, on the cell surface and along the phagosome membrane. It is, however, very weak in resident macrophages and increases after stimulation or activation. The second kind of bactericidal property corresponds to cationic proteins located in lysosomes. After fusion between lysosomes and phagosomes, they contribute to microbe killing by permeabilizing microbe envelopes. Lysosomes, which contain diverse acid hydrolases and are responsible for the degradation of ingested material, play a crucial role in macrophage endocytic activity. Their number increases in parallel with endocytic activity during macrophage differentiation and is particularly high after ingestion of degradable material. Contrary to polymorphonuclear leukocytes, macrophage is very poor in granules containing peroxidase. The latter, which are rather abundant in monocytes, disappear during macrophage maturation. They do not seem thus to be implicated in macrophage microbicidal activity. Endocytosis is accompanied by rapid and intense exchanges between the different membrane compartments of the cell (plasma membrane, pinosomes or phagosomes, endosomes, lysosomes, Golgi apparatus, etc.). These exchanges seem to occur by transitory fusions between vesicles coming from different compartments, rapidly followed by their recycling to their original compartment. This system of membrane shuttle has been clearly observed after formation of phagosomes or pinosomes in which the internalized plasma membrane is recycled back to the cell surface within a few minutes after their formation. This membrane traffic is especially intense in macrophages, the endocytic activity of which is very high, but it also exists in all cell types.(ABSTRACT TRUNCATED AT 400 WORDS)
36 citations
TL;DR: Results show that oxidative metabolism in amebae has some striking similarities to the respiratory burst oxidase of neutrophils.
Abstract: When amebae were incubated with latex beads, cyanide-insensitive oxygen consumption increased nearly two-fold. This cyanide-insensitive respiration was inhibited by salicylhydroxamate. Furthermore, cell fractionation studies revealed a localization for a portion of the NAD(P)H oxidase activity in phagolysosomes. The presence of low concentrations of divalent metal during fractionation resulted in an increased yield of oxidative activity in the phagolysosome fraction. In addition, the phagolysosome membrane was enriched about two-fold in a b-type cytochrome. These results show that oxidative metabolism in amebae has some striking similarities to the respiratory burst oxidase of neutrophils.
19 citations
TL;DR: It is found that TMPase is a lysosomal enzyme of mammalian macrophages and may take part in the degradation of phagocytosed materials, particularly microorganisms which contain inorganic polyphosphates and metaphosphates.
15 citations
TL;DR: The actin which is newly associated with the surface membrane of the phagocytosing cells was more susceptible to removal by detergent than the membrane-associated actin in resting cells and it was also noted that the F-actin associated with phagosomes was readily disrupted by Detergent.
Abstract: In the analysis of highly purified surface membrane from both resting and phagocytosing neutrophils an increase in the surface membrane associated actin has been demonstrated. This change at the cell periphery is associated with a coincident increase in the F-actin content of the cells following stimulation of the cells by exposure to opsonized Oil Red O droplets. The actin which is newly associated with the surface membrane of the phagocytosing cells was more susceptible to removal by detergent than the membrane-associated actin in resting cells and it was also noted that the F-actin associated with phagosomes was readily disrupted by detergent. A redistribution of the surface membrane glycoprotein 5'-nucleotidase was observed during phagocytosis, but no change in distribution of a 125I-labelled Lens culinaris lectin was observed during the entire phagocytic process.
11 citations
TL;DR: Both in vivo and in vitro, Mycobacterium marinum organisms were found to multiply within phagolysosomes of murine macrophages, and it thus appears that M. Marinum are neither killed nor inhibited from multiplying by lysosomal enzymes.
Abstract: Both in vivo and in vitro, Mycobacterium marinum organisms were found to multiply within phagolysosomes of murine macrophages. It thus appears that M. marinum are neither killed nor inhibited from multiplying by lysosomal enzymes.
9 citations
TL;DR: The reaction of the contractile vacuole of Amoeba proteus to single and multiple phagocytosis under controlled conditions has been studied and some form of active transport is implied.
Abstract: The reaction of the contractile vacuole of Amoeba proteus to single and multiple phagocytosis under controlled conditions has been studied. Fluid intake into the cytoplasm from the phagosomes induces secretion by the contractile vacuole of equivalent excess volumes:. Vacuolar response is rapid (200 sec) and may be initiated by increases of protoplasmic hydration of as little as 1%. Cytoplasmic uptake of fluid from the phagosome can occur against an osmotic gradient; thus some form of active transport is implied.
7 citations
01 Jan 1985
TL;DR: Fusion-promoting lipophilic secondary and tertiary amines are particularly interesting since normal mouse macrophages infected with M.tuberculosis suppress the infection if treated with these bases (e.g., chloroquine, tributylamine, dibenzylamine) (7).
Abstract: After phagocytosis by cultured macrophages, certain species of pathogenic microorganisms evade exposure to the contents of secondary lysosomes by ensuring the nonfusion of their phagosomes with these organelles. Circumstantial evidence suggests that this nonfusion is the result of inhibitory influences by the microorganisms, exerted from within the phagosomes. Examples are Mycobacterium tuberculosis (1), Toxoplasma gondii (2), Chlamydia psittaci (3) and Legionella pneumophila (4). Inhibition or enhancement of phagosome-lysosome (P-L) fusion in cultured macrophages provided with a suitable phagocytic target, e.g., yeast cells can be induced also by certain lysosomotropic chemical agents. These agents are mainly polyanions (inhibitors) and nitrogenous bases, notably amine weak bases (e.g., chloroquine, an enhancer, and ammonium chloride, an inhibitor) (5–9). Fusion-promoting lipophilic secondary and tertiary amines are particularly interesting since normal mouse macrophages infected with M.tuberculosis suppress the infection if treated with these bases (e.g., chloroquine, tributylamine, dibenzylamine) (7).
4 citations
Journal Article•
TL;DR: Indirect fluorescent, radio- and peroxidase immunobinding assays with intact and methanol-permeabilized cells confirmed that antigens PL-1 and PL-4 were exclusively intracellular and that well-washed phagolysosomes bound both antibodies.
Abstract: Monoclonal antibodies were prepared to study the cytoplasmic face of latex phagolysosomes isolated from thioglycollate-elicited mouse peritoneal macrophages. Phagolysosomes obtained by sucrose flotation contained latent beta-glucuronidase activity and tightly associated cellular proteins and glycoproteins. Fluorescence-activated cell sorter analysis, scanning and transmission electron microscopy showed that the particle preparation contained greater than 98% monomers and dimers, invested with a smooth layer of membrane and minimally contaminated with cytoplasmic adhesions. Sera for immunized rats bound preferentially to isolated phagolysosomes rather than intact cells and monoclonal antibodies PL-1 and PL-4 were isolated on this basis. Indirect fluorescent, radio- and peroxidase immunobinding assays with intact and methanol-permeabilized cells confirmed that antigens PL-1 and PL-4 were exclusively intracellular and that well-washed phagolysosomes bound both antibodies. These antigens were found in a variety of cells from several species and in macrophages not fed latex. Although the PL-1 antigen could not be immunoprecipitated, intracellular staining was characteristic of intermediate filament distribution, that is, it was in the form of a fine intersecting network, which collapsed, reversibly, in a rim round the nucleus upon treatment with colcemid. The staining pattern was undetectable in cells 1 h after adherence to a substratum, but gradually appeared after 6–12 h. The PL-4 antibody has been shown elsewhere to define a Ca2+-binding protein of approximately 20 000 molecular weight, which is phosphorylated during phagocytosis. This antibody stained stress fibres and revealed a widespread punctate distribution of antigen within cells at all stages after adhesion. The nature of the association between these intracellular antigens and phagolysosomes and their possible role in phagocytosis are not known.
TL;DR: The evidence suggests that monoclonal antibodies against capped membrane are useful probes for analysing functional cytoskeleton domains; and the 41.5K-dalton component is probably involved in the phagocytosis.
Abstract: The formation of the ligand-receptor complex on cell surface causes a tight association of microfilaments with the plasma membrane Since capping is an energy-dependent process which involves contractile proteins, it seemed reasonable to look for physiologically important cytoskeletal elements in regions of the capped membrane We isolated the capped membrane ofDictyostelium induced with lentil lectin, separated the proteins by electrophoresis, and prepared monoclonal antibodies against proteins extracted from the gel These antibodies were reactive to distinct antigens involved in the cytoskeleton-membrane complex, as observed by improved immunofluorescence (“agar-overlay” technique) We further characterized one of these antibodies (DCC-32) (DCC:Dictyostelium cortical cytoskeleton), and found that it was reactive to a new cytoskeletal element of 415 K (pI 1065) according to a two-dimensional Western blotting Immunoelectron microscopy showed this antigen to be localized in the cortical cytoskeleton associated with the plasma membrane and at the cytoplasmic side of the phagosome membrane The evidence suggests that monoclonal antibodies against capped membrane are useful probes for analysing functional cytoskeleton domains; and the 415K-dalton component is probably involved in the phagocytosis
01 Jan 1985
TL;DR: The unique modified endocytic vacuole occupied by Toxoplasma is characterized by an accumulation of membrane-like tubules which may contribute to the toxoplasmacidal responses of these cells.
Abstract: Toxoplasma gondii is a protozoan parasite which resides in modified endocytic compartments of host cells. Extensive intracellular replication and host cell lysis leads to acute toxoplasmosis which is eventually limited by the host immune response to a chronic state of infection. The features of this unique intracellular compartment, which enables Toxoplasma to survive in macrophages, are reported here. Although Toxoplasma survives in normal macrophages, activated macrophages from immune animals rapidly killed Toxoplasma by production of oxygen radicals and intermediates generated during parasite invasion. In addition, activated macrophages inhibited Toxoplasma growth by an oxygen-independent mechanism. Qualitative and quantitative features of oxygen intermediate detoxifying enzymes, catalase and superoxide dismutase, were described from two strains of Toxoplasma. These enzymes did not appear to be the basis for differences in strain virulence, but may contribute to intracellular survival in normal macrophages. A newly recognized microbicidal mechanism involves the rapid acidification during the formation of endocytic compartments. Live Toxoplasma entered into modified phagocytic vacuoles in normal macrophages that do not show characteristic acidification, but remain at near neutral pH. In contrast, the enhanced acidificaton capacity of activated macrophages and of normal macophages in the presence of specific antibody may contribute to the toxoplasmacidal responses of these cells. The unique modified endocytic vacuole occupied by Toxoplasma is characterized by an accumulation of membrane-like tubules which may