Tracing iron and transferrin in the macrophage by visual means.

The purpose of the study was to investigate the movement of iron and transferrin in the macrophage using light and electron microscopy. First, depicted here are the phagocytosis of antibody sensitized murine red cells by the murine bone marrow derived macrophage and the formation of red cell phagosomes. Second, we show the fusion of the lysosomes with the red cell phagosome to form a lysophagosome and the lysis of the red cell using acid phosphatase as a lysosome marker.

Intracellular differentiation of Leishmania amazonensis promastigotes to amastigotes: presence of megasomes, cysteine proteinase activity and susceptibility to leucine-methyl ester.

Intracellular differentiation of Leishmania promastigotes to amastigotes is a critical step in the establishment of infection. In this report three related features of mexicana subspecies amastigotes were used to follow the differentiation of the parasites within macrophages. Early after infection, (a) parasites did not contain ultrastructurally recognizable megasomes, (b) cysteine proteinase activity of parasite lysates was not detected in gelatin-containing acrylamide gels, and (c) parasites were essentially resistant to L-leucine-methyl ester (Leu-OMe).

HIV-1 infection of lung alveolar fibroblasts and macrophages in humans.

We have studied the infected cell populations in the lungs of four human immunodeficiency virus type 1 (HIV-1) seropositive patients suffering from lymphocytic alveolitis or lymphocytic interstitial pneumonitis. Adherent cells were obtained by bronchoalveolar lavage (BAL) and were analyzed by various technical approaches. The cells considered here were alveolar macrophages and fibroblasts, and could be clearly identified morphologically and by the expression of specific cell-surface markers using monoclonal antibodies.

Modified lysosomal compartment as carrier of slowly and non-degradable tracers in macrophages.

A previous immunocytochemical study of macrophages infected with Bacillus subtilis showed that a cell wall antigen could be detected for several days in a population of small vesicles randomly distributed within the cells and apparently distinct from perinuclear lysosomes. These observations suggested the possibility that these vesicles might constitute a "storage" compartment for non-degradable compounds. In the present report we compared in pulse-chase experiments the location and fate of a series of degradable and non-degradable pinocytic tracers within the macrophages.

Contribution of electron microscopy in the study of the interactions between pathogenic bacteria and their host cells.

Our knowledge on the functional anatomy of bacteria is based on the electron microscopic (EM) studies performed during the last forty years. Most pathogenic properties however cannot be visualized in EM because they are not related to defined structures. In contrast, EM studies have provided important data on the behaviour of pathogenic bacteria in their host cells.

Immunolabelling of Shiga toxin in macrophages infected with Shigella dysenteriae 1.

Immunolabelling of Shiga toxin in macrophages infected with a non-invasive Shigella dysenteriae 1 isolate showed that bacteria remained alive for 3 h after ingestion within the phagocytic vacuole and synthesized Shiga toxin. The normal process of toxin secretion was, however, impaired by the phagosomal environment and toxin molecules accumulated within the bacterial cytoplasm.

Improved sectioning and ultrastructure of bacteria and animal cells embedded in Lowicryl.

Lowicryl K4M-embedded Gram-positive and Gram-negative bacteria have a tendency to separate between the cell surface and the resin. This often leads to distortion of bacteria and more especially of mycobacteria. We describe attempts made to overcome this technical problem.

Intracellular and cell-to-cell spread of Listeria monocytogenes involves interaction with F-actin in the enterocytelike cell line Caco-2.

Listeria monocytogenes penetrates and multiplies within professional phagocytes and other cells such as the Caco-2 human enterocytelike cell line. Listeriolysin O, a membrane-damaging cytotoxin accounts for intracellular multiplication through lysis of the membrane-bound phagocytic vacuole. This work demonstrates that once released within the cytosol, L.

Analysis of the subcellular location of pullulanase produced by Escherichia coli carrying the pulA gene from Klebsiella pneumoniae strain UNF5023.

Three different techniques, protease accessibility, cell fractionation and in situ immunocytochemistry, were used to study the location of the lipoprotein pullulanase produced by Escherichia coli K12 carrying the cloned pullulanase structural gene (pulA) from Klebsiella pneumoniae, with or without the K. pneumoniae genes required to transport pullulanase to the cell surface (secretion-competent and secretion-incompetent, respectively). Pullulanase produced by secretion-competent strains could be slowly but quantitatively released into the medium by growing the cells in medium containing pronase.

Immunodetection of lipopolysaccharide in macrophages during the processing of non invasive Shigella dysenteriae.

The location of lipopolysaccharide (LPS) was studied by immunofluorescence and immunoelectron microscopy in macrophages infected with a non-invasive Shigella dysenteriae 1 strain. Bacterial degradation began only 3 h after the end of infection. The first visible sign of degradation was detected by immunogold labelling at the level of LPS which detached from the bacterial surface and was transferred to the perinuclear lysosomes.

Heterologous protein export in Escherichia coli: influence of bacterial signal peptides on the export of human interleukin 1 beta.

Expression plasmids carrying the coding sequence of mature human interleukin 1 beta (IL 1 beta) linked either to a Met start codon, or fused to different efficient Escherichia coli secretion signal sequences, have been constructed. In the latter case, we used signal peptides derived either from an outer membrane protein (OmpA) or from a periplasmic protein (PhoA). The synthesis of IL1 beta from these fusions was investigated in an otherwise strictly isogenic context using identical conditions of derepression and culture media.

Antibodies against synthetic peptides and the topology of LamB, an outer membrane protein from Escherichia coli K12.

LamB, an outer membrane protein from Escherichia coli K12, is involved in the transport of maltose and maltodextrins across the outer membrane and constitutes a receptor for a number of bacteriophages. A recent folding model proposes that LamB spans the outer membrane through a number of transmembranous segments separated by regions exposed either to the cell exterior or to the periplasm. This model is essentially based on predictions of structure and genetic arguments relying on the hypothesis that the mutations studied did not alter the folding of the protein.

Megasomes as the targets of leucine methyl ester in Leishmania amazonensis amastigotes.

Certain L-amino acid esters, such as L-leucine methyl ester (Leu-OMe), can kill intracellular and isolated Leishmania amazonensis amastigotes. Killing appears to involve ester trapping and hydrolysis within an acidified parasite compartment (M. Rabinovitch and S.

Leishmania amazonensis: acidic organelles in amastigotes.

Leishmania amastigotes are intracellular protozoan parasites which exclusively invade cells of the macrophage series and multiply within phagolysosomes. Recent studies showed that intracellular and isolated amastigotes of L. amazonesis are killed by amino acid esters which appear to be trapped within as yet unidentified, possibly acidified, "lysosome-like" parasite compartments and cleaved by hydrolytic enzyme(s) (M.

Endocytic membrane traffic with respect to phagosomes in macrophages infected with non-pathogenic bacteria: phagosomal membrane acquires the same composition as lysosomal membrane.

A morphometric analysis was made to study membrane traffic in bone marrow-derived macrophages, containing phagosomes with partially degraded Bacillus subtilis. Cell surface glycoproteins, labeled with radioactive galactose by terminal glycosylation, provided a covalent autoradiographic membrane marker. Membrane compartments were characterized in terms of cytochemical staining for horseradish peroxidase taken up by receptor-mediated endocytosis.

Bacterial antigen immunolabeling in macrophages after phagocytosis and degradation of Bacillus subtilis.

After phagocytosis of Bacillus subtilis 168 by bone marrow-derived macrophages, the intracellular pathway followed by different antigens was studied by immunofluorescence and immunoelectron microscopy. Three different rabbit antisera were used: (i) an antiserum to B. subtilis whole cells mainly recognizing the cell wall constituents, (ii) an antiserum to teichoic acid, and (iii) an antiserum to peptidoglycan recognizing the disaccharide tetrapeptide molecules resulting from peptidoglycan degradation.

Induction of a stable morphological change in Propionibacterium freudenreichii.

When cells of Propionibacterium freudenreichii were incubated under fasting conditions and then plated in the presence of an inhibitor of protein synthesis, a variable but significant (greater than 10(-2) fraction of the population changed their morphology from rod to sphere, with a considerable thickening of the cell wall. This change was accompanied by metabolic and antibiotic-resistance modifications, including the synthesis of at least one new enzyme (alpha-glucosidase), and by the simultaneous appearance of several new species of DNA, presumably plasmids. The round cells grew faster than the parent strain and maintained their morphology indefinitely when propagated on complex medium containing glucose as the main carbon source.

Contribution of new cryomethods to a better knowledge of bacterial anatomy.

Electron microscopy has largely contributed to the study of bacterial anatomy. However, as varied alterations can occur during cell preparation, at the level of cell structure and at the molecular level, it is difficult to know to what extent electron micrographs correspond to the true appearance of the living state. The recent development of cryomethods which avoid some of the alterations which may occur during conventional fixation and embedding procedures, has shed new light on bacterial anatomy.

[A new sensitivity test for substances used as antiseptics or disinfectants. I. Biological studies].

In this first article, the authors present a fast method of determining the bactericide and bacteriostatic power of the germs, based on the disparition of the succino-dehydrogenase activity of a bacterium, objectified by a neotetrazolium chloride reaction. This reaction allows a colorimetric qualitative and quantitative determination of the activity of a preparation and the duration of its contact with the bacteria. This method allows a study in electronic microscopy which will be published in a second article.

[Ultrastructural changes induced by antiseptics in various bacterial species].

Due to the succinate-neotetrazolium technique, the authors were able to photograph with electronic microscopy, for the first time, different antiseptics, including "Javelle Water", attacking bacteria.

Leishmania mexicana: a cytochemical and quantitative study of lysosomal enzymes in infected rat bone marrow-derived macrophages.

The cellular localization and activity of the lysosomal enzymes acid phosphatase, trimetaphosphatase, and arylsulfatase were studied in rat bone marrow-derived macrophages infected with Leishmania mexicana amazonensis amastigotes. The specific activity of acid phosphatase normalized for protein content was similar in normal macrophages and in isolated amastigotes, whereas the latter were markedly deficient in trimetaphosphatase and arylsulfatase activities. It is thus likely that trimetaphosphatase and arylsulfatase activities detected in infected macrophages were of host cell origin.