scholarly journals THE FATE OF BACTERIA WITHIN PHAGOCYTIC CELLS

1964 ◽  
Vol 120 (5) ◽  
pp. 869-883 ◽  
Author(s):  
Zanvil A. Cohn
Keyword(s):  
Alveolar Macrophages ◽  
Peritoneal Macrophages ◽  
Immune Serum ◽  
Soluble Form ◽  
Phagocytic Cells ◽  
E Coli ◽  
Heat Stable ◽  
Immune Globulins

The fate of a heat-stable Escherichia coli agglutinogen within three types of rabbit phagocytic cells was examined. A system is described whereby quantitative ingestion of viable E. coli by suspensions of PMN leucocytes, BCG-induced alveolar macrophages, and oil-induced peritoneal macrophages took place in vitro. After various periods of intracellular residence aliquots were injected intraperitoneally into NCS mice and the resulting agglutinins assayed. The loss of immunogenicity within phagocytes was estimated by comparison with a dose-response titration prepared with bacteria alone. Under these conditions no increase in immunogenic mass occurred in vivo or in vitro when viable organisms were employed. PMN leucocytes and alveolar macrophages destroyed the majority of the immunogen within 2 hours of intracellular residence. In contrast, the immunogenicity of E. coli was maintained within peritoneal macrophages for periods up to 5 hours. The use of heat-killed bacilli or specific immune serum did not significantly influence the intracellular fate of the immunogen. Residual immunogenicity was associated with a particle having the same centrifugal properties as the intact organism and essentially none was released in a soluble form. Intracellular residence within phagocytic cells did not influence the resulting temporal sequence of antibody formation nor the proportions of mercaptoethanol-sensitive and resistant immune globulins.

scholarly journals Full Capacity of Recombinant Escherichia coliHeat-Stable Enterotoxin Fusion Proteins for Extracellular Secretion, Antigenicity, Disulfide Bond Formation, and Activity

2000 ◽  
Vol 68 (7) ◽  
pp. 4064-4074 ◽  
Author(s):  
Isabelle Batisson ◽  
Maurice Der Vartanian ◽  
Brigitte Gaillard-Martinie ◽  
Michel Contrepois
Keyword(s):  
Disulfide Bonds ◽  
Secretory Pathway ◽  
Biological Properties ◽  
Leader Peptide ◽  
Dependent Manner ◽  
Reporter Protein ◽  
E Coli ◽  
Heat Stable

ABSTRACT We have successfully used the major subunit ClpG ofEscherichia coli CS31A fimbriae as an antigenic and immunogenic exposure-delivery vector for various heterologous peptides varying in nature and length. However, the ability of ClpG as a carrier to maintain in vitro and in vivo the native biological properties of passenger peptide has not yet been reported. To address this possibility, we genetically fused peptides containing all or part of the E. coli human heat-stable enterotoxin (STh) sequence to the amino or carboxyl ends of ClpG. Using antibodies to the ClpG and STh portions for detecting the hybrids; AMS (4-acetamido-4′-maleimidylstilbene-2,2′-disulfonate), a potent free thiol-trapping reagent, for determining the redox state of STh in the fusion; and the suckling mouse assay for enterotoxicity, we demonstrated that all ClpG-STh proteins were secreted in vitro and in vivo outside the E. coli cells in a heat-stable active oxidized (disulfide-bonded) form. Indeed, in contrast to many earlier studies, blocking the natural NH2 or COOH extremities of STh had, in all cases, no drastic effect on cell release and toxin activity. Only antigenicity of STh C-terminally extended with ClpG was strongly affected in a conformation-dependent manner. These results suggest that the STh activity was not altered by the chimeric structure, and therefore that, like the natural toxin, STh in the fusion had a spatial structure flexible enough to be compatible with secretion and enterotoxicity (folding and STh receptor recognition). Our study also indicates that disulfide bonds were essential for enterotoxicity but not for release, that spontaneous oxidation by molecular oxygen occurred in vitro in the medium, and that the E. coli cell-bound toxin activity in vivo resulted from an effective export processing of hybrids and not cell lysis. None of the ClpG-STh subunits formed hybrid CS31A-STh fimbriae at the cell surface ofE. coli, and a strong decrease in the toxin activity was observed in the absence of CS31A helper proteins. In fact, chimeras translocated across the outer membrane as a free folded monomer once they were guided into the periplasm by the ClpG leader peptide through the CS31A-dependent secretory pathway. In summary, ClpG appears highly attractive as a carrier reporter protein for basic and applied research through the engineering of E. coli for culture supernatant delivery of an active cysteine-containing protein, such as the heat-stable enterotoxin.

scholarly journals Role of Myeloid Tet Methylcytosine Dioxygenase 2 in Pulmonary and Peritoneal Inflammation Induced by Lipopolysaccharide and Peritonitis Induced by Escherichia coli

Cells ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 82
Author(s):  
Wanhai Qin ◽  
Xanthe Brands ◽  
Hisatake Matsumoto ◽  
Joe M. Butler ◽  
Cornelis van’t Veer ◽  
...  
Keyword(s):  
Peritoneal Lavage ◽  
Lavage Fluid ◽  
Peritoneal Macrophages ◽  
Histone Deacetylation ◽  
Peritoneal Lavage Fluid ◽  
E Coli ◽  
Peritoneal Inflammation ◽  
Enhanced Production

Tet methylcytosine dioxygenase 2 (Tet2) mediates demethylation of DNA. We here sought to determine the expression and function of Tet2 in macrophages upon exposure to lipopolysaccharide (LPS), and in the host response to LPS induced lung and peritoneal inflammation, and during Escherichia (E.) coli induced peritonitis. LPS induced Tet2 expression in mouse macrophages and human monocytes in vitro, as well as in human alveolar macrophages after bronchial instillation in vivo. Bone marrow-derived macrophages from myeloid Tet2 deficient (Tet2fl/flLysMCre) mice displayed enhanced production of IL-1β, IL-6 and CXCL1 upon stimulation with several Toll-like receptor agonists; similar results were obtained with LPS stimulated alveolar and peritoneal macrophages. Histone deacetylation was involved in the effect of Tet2 on IL-6 production, whilst methylation at the Il6 promoter was not altered by Tet2 deficiency. Tet2fl/flLysMCre mice showed higher IL-6 and TNF levels in bronchoalveolar and peritoneal lavage fluid after intranasal and intraperitoneal LPS administration, respectively, whilst other inflammatory responses were unaltered. E. coli induced stronger production of IL-1β and IL-6 by Tet2 deficient peritoneal macrophages but not in peritoneal lavage fluid of Tet2fl/flLysMCre mice after in vivo intraperitoneal infection. Tet2fl/flLysMCre mice displayed enhanced bacterial growth during E. coli peritonitis, which was associated with a reduced capacity of Tet2fl/flLysMCre peritoneal macrophages to inhibit the growth of E. coli in vitro. Collectively, these data suggest that Tet2 is involved in the regulation of macrophage functions triggered by LPS and during E. coli infection.

In vitro and in vivo Evaluation of 111In-labeled E. coli Heat-Stable Enterotoxin Analogs for Specific Targeting of Human Breast Cancers

2006 ◽  
Vol 98 (1) ◽  
pp. 7-15 ◽  
Author(s):  
Michael F. Giblin ◽  
Hariprasad Gali ◽  
Gary L. Sieckman ◽  
Nellie K. Owen ◽  
Timothy J. Hoffman ◽  
...  
Keyword(s):  
Breast Cancers ◽  
Human Breast ◽  
In Vivo Evaluation ◽  
E Coli ◽  
Specific Targeting ◽  
Heat Stable

scholarly journals An Atypical KdpD Homologue from the Cyanobacterium Anabaena sp. Strain L-31: Cloning, In Vivo Expression, and Interaction with Escherichia coli KdpD-CTD

2005 ◽  
Vol 187 (14) ◽  
pp. 4921-4927 ◽  
Author(s):  
Anand Ballal ◽  
Marc Bramkamp ◽  
Hema Rajaram ◽  
Petra Zimmann ◽  
Shree Kumar Apte ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Response Regulator ◽  
Nitrilotriacetic Acid ◽  
Soluble Form ◽  
Filamentous Cyanobacterium ◽  
E Coli ◽  
Transmembrane Segments ◽  
Terminal Domain

ABSTRACT The kdpFABC operon of Escherichia coli, coding for the high-affinity K+ transport system KdpFABC, is transcriptionally regulated by the products of the adjacently located kdpDE genes. The KdpD protein is a membrane-bound sensor kinase consisting of a large N-terminal domain and a C-terminal transmitter domain interconnected by four transmembrane segments (the transmembrane segments together with the C-terminal transmitter domain of KdpD are referred to as CTD), while KdpE is a cytosolic response regulator. We have cloned and sequenced the kdp operon from a nitrogen-fixing, filamentous cyanobacterium, Anabaena sp. strain L-31 (GenBank accession. number AF213466 ). The kdpABC genes are similar in size to those of E. coli, but the kdpD gene is short (coding only for 365 amino acids), showing homology only to the N-terminal domain of E. coli KdpD. A kdpE-like gene is absent in the vicinity of this operon. Anabaena KdpD with six C-terminal histidines was overproduced in E. coli and purified by Ni2+-nitrilotriacetic acid affinity chromatography. With antisera raised against the purified Anabaena KdpD, the protein was detected in Anabaena sp. strain L-31 membranes. The membrane-associated or soluble form of the Anabaena KdpD(6His) could be photoaffinity labeled with the ATP analog 8-azido-ATP, indicating the presence of an ATP binding site. The coproduction of Anabaena KdpD with E. coli KdpD-CTD decreased E. coli kdpFABC expression in response to K+ limitation in vivo relative to the wild-type KdpD-CTD protein. In vitro experiments revealed that the kinase activity of the E. coli KdpD-CTD was unaffected, but its phosphatase activity increased in the presence of Anabaena KdpD(6His). To our knowledge this is the first report where a heterologous N-terminal domain (Anabaena KdpD) is shown to affect in trans KdpD-CTD (E. coli) activity, which is just opposite to that observed for the KdpD-N-terminal domain of E. coli.

In vitro and in vivo evaluation of 177Lu- and 90Y-labeled E. coli heat-stable enterotoxin for specific targeting of uroguanylin receptors on human colon cancers

2006 ◽  
Vol 33 (4) ◽  
pp. 481-488 ◽  
Author(s):  
Michael F. Giblin ◽  
Gary L. Sieckman ◽  
Tiffani D. Shelton ◽  
Timothy J. Hoffman ◽  
Leonard R. Forte ◽  
...  
Keyword(s):  
Human Colon ◽  
In Vivo Evaluation ◽  
E Coli ◽  
Specific Targeting ◽  
Colon Cancers ◽  
Heat Stable

Activated macrophages in highly irradiated cercariae-induced immunity toSchistosoma japonicumin rats

Parasitology ◽  
1991 ◽  
Vol 102 (1) ◽  
pp. 65-72
Author(s):  
C. Xu ◽  
S. Xu
Keyword(s):  
Alveolar Macrophages ◽  
Sheep Erythrocyte ◽  
Peritoneal Macrophages ◽  
Rat Serum ◽  
Sheep Erythrocytes ◽  
Proteose Peptone ◽  
Normal Rat ◽  
Induced Immunity

SUMMARYThe results of studies on the schistosomulicidal activity of activated peritoneal and alveolar macrophages (pMø and aMø) from rats immunized with highly irradiated (50 krad.)Schistosoma japonicumcercariae are reported. The authors have examined the activation of these macrophages in terms of spreading, adhesion and ingestion of sheep erythrocytes and pinocytosis of horse-radish peroxidase. Using three criteria, peritoneal macrophages and alveolar macrophages from immunized rats and from rats intraperitoneally injected with BCG were significantly more active than those from normal rats or rats stimulated with 10% proteose-peptone or 1% sodium thioglycolate. A significantly higher percentage of adhesion and ingestion was obtained with the sheep erythrocytes that were co-opsonized by heat-inactivated rat anti-sheep erythrocyte serum and fresh normal rat serum. Schistosomulicidal effects were observed with macrophages from irradiated cercariae-immunized rats in two activation systems:in vitroactivation in the presence of macrophage-activating factor (MAF), andin vivoactivation by the intraperitoneal challenge with sonicated cercarial antigens.

Rat alveolar macrophages express preprotachykinin gene-I mRNA-encoding tachykinins

1997 ◽  
Vol 273 (5) ◽  
pp. L1073-L1081 ◽  
Author(s):  
Cheryl R. Killingsworth ◽  
Stephanie A. Shore ◽  
Francesca Alessandrini ◽  
Richard D. Dey ◽  
Joseph D. Paulauskis
Keyword(s):  
Alveolar Macrophages ◽  
Peritoneal Macrophages ◽  
Northern Analysis ◽  
Neurokinin A ◽  
Polymerase Chain ◽  
Preprotachykinin Gene ◽  
Lps Treatment

Although the tachykinins substance P (SP) and neurokinin A have been largely localized to neurons, eosinophils have also been shown to express these peptides. Our aim was to determine whether rat alveolar macrophages (AM) express preprotachykinin gene-I (PPT-I) mRNA that encodes these tachykinins and to examine expression during inflammation. PPT-I mRNA was detected by reverse transcription (RT)-polymerase chain reaction (PCR) in AM and brain (control) but not in peritoneal macrophages. Northern analysis showed that PPT-I mRNA was induced two- to fourfold by in vivo treatment of rats with intratracheal lipopolysaccharide (LPS) and in vitro after 4 h of exposure to LPS. This increase was inhibited by dexamethasone. In situ RT-PCR and immunocytochemistry further confirmed that AM express PPT-I mRNA and SP-like immunoreactivity, respectively, which was enhanced by LPS treatment. A 1.3-kb transcript consistent with PPT-I mRNA was detected by Northern analysis of bronchoalveolar lavage neutrophils. Therefore, rat AM express PPT-I mRNA that is upregulated in AM by LPS and is attenuated by dexamethasone. PPT-I mRNA was also detected in lung neutrophils.

scholarly journals THE ROLE OF OPSONINS IN NON-SPECIFIC IMMUNITY

1960 ◽  
Vol 111 (1) ◽  
pp. 137-144 ◽  
Author(s):  
Derrick Rowley
Keyword(s):  
Normal Serum ◽  
Phagocytic Cells ◽  
E Coli ◽  
Specific Immunity ◽  
Viable Bacteria ◽  
Bacterial Lipopolysaccharides ◽  
Transport Of Bacteria

A study of the recoveries of radioactivity, and of viable bacteria following injection of P32-labelled E. coli into the mouse peritoneum, has indicated that the rapid decrease in viable bacteria which occurs is largely due to peritoneal events, and not to the transport of bacteria elsewhere. The serum from mice given lipopolysaccharides 48 hours previously, when used to pretreat bacteria before intraperitoneal injection, was found to stimulate phagocytosis to a greater extent than did pretreatment with normal serum. In addition, macrophages themselves were found to be affected by contact with lipopolysaccharides, either in vivo or in vitro in such a way as to promote their phagocytic abilities. It is suggested that the provocation of non-specific immunity by bacterial lipopolysaccharides involves two facets at least; firstly, an increase in the opsonic capacity of the serum, and secondly an increase in the inherent capacity of phagocytic cells to perform this function.

scholarly journals Release of Fibrinolytic Enzymes by Macrophages in Response to Soluble Fibrin

1977 ◽  
Author(s):  
L. A. Sherman ◽  
J. Lee ◽  
C. C. Stewart
Keyword(s):  
Plasminogen Activator ◽  
Proteolytic Enzymes ◽  
Peritoneal Macrophages ◽  
Phagocytic Cells ◽  
Fibrinolytic Enzymes ◽  
Soluble Fibrin ◽  
The Media ◽  
Free Media

In previous data (J. Exp. Med. 147:76,1977), we have demonstrated that soluble fibrin/fibrinogen complexes are bound to the plasma membrane of guinea pig peritoneal macrophages. This binding is largely irreversible and is not a result of phagocytosis. We have extended our studies to examine the response in vitro of peritoneal macrophages to soluble fibrin/fibrinogen complexes. Unstimulated mouse macrophages were collected by peritoneal lavage and 5–60 μg of soluble fibrin/fibrinogen complexes placed into tissue culture dishes containing the unstimulated cells. Aliquots of the media were collected at 24, 48 and 72 hours. The cell-free media contained increasing amounts both of plasminogen activator and an enzymatic activity which resulted in fibrin and fibrinogen proteolysis independent of the amount of plasmingoen present. The major proteolytic activity was due to the non-plasminogen dependent enzyme. Similar enzymes were released from peritoneal macrophages stimulated in vivo. The plasminogen activator enzyme had a low molecular weight comparable to that previously reported by Unkeless et al, with in vivo stimulation. Other coagulation moieties, such as plasmin and α-2 macroglobulin plasmin complexes did not result in release of the macrophage proteolytic enzymes. The results suggest that the previously described release of fibrinolytic enzymes after thioglycolate injections, may also result from the more pathophysiological stimulation by soluble fibrin/fibrinogen complexes. Release of these enzymes from phagocytic cells may be important, not only in blood clearance of soluble fibrin/fibrinogen complexes, but as part of thrombus reabsorption and wound healing.

scholarly journals Effects of immunization and anticoagulation on the development of experimental Escherichia coli endocarditis

1980 ◽  
Vol 28 (2) ◽  
pp. 325-330
Author(s):  
L Thörig ◽  
J Thompson ◽  
R van Furth
Keyword(s):  
Escherichia Coli ◽  
Peritoneal Macrophages ◽  
Immune Serum ◽  
Bacterial Endocarditis ◽  
Intracellular Killing ◽  
Opsonic Activity ◽  
Specific Antibodies ◽  
E Coli ◽  
Number Of Bacteria

The effects of immunization and anticoagulation in experimental Escherichia coli endocarditis were studied. Immunization of rabbits with E. coli resulted in the development of specific agglutinating and opsonic activity of the serum, but not in bactericidal activity. These antibody activities also developed in nonimmunized rabbits during the course of bactericidal endocarditis. Immune serum promoted phagocytosis in vitro but did not enhance intracellular killing of E. coli by elicited rabbit peritoneal macrophages. The presence of specific antibodies in rabbits after immunization had no effect on the induction or course of E. coli infection of endocardial vegetations. Anticoagulation was found to affect the induction of the infection. In anticoagulated rabbits, larger bacterial inocula were needed to induce an infection, but in animals with bacterial endocarditis the number of bacteria in the vegetations did not differ significantly from that of the control animals.

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