scholarly journals Monoclonal antibody with hemagglutination, immobilization, and neutralization activities defines an immunodominant, 47,000 mol wt, surface-exposed immunogen of Treponema pallidum (Nichols).

1984 ◽  
Vol 160 (5) ◽  
pp. 1404-1420 ◽  
Author(s):  
S A Jones ◽  
K S Marchitto ◽  
J N Miller ◽  
M V Norgard
Keyword(s):  
Treponema Pallidum ◽  
Immune Serum ◽  
Host Cells ◽  
Rabbit Serum ◽  
Functional Activities ◽  
Molecular Weights ◽  
Rabbit Igg ◽  
Major Surface

Radioimmunoprecipitation (RIP) analyses performed on 125I-surface-labeled Treponema pallidum cells using various immune sera revealed the presence of six major surface antigens (immunogens) with apparent molecular weights of 47 K, 36 K, 34 K, 32 K, 29 K, and 13 K. Among these, the 47 K surface antigen was most abundant. Radioimmunoprecipitation assays using 125I-labeled T. phagedenis biotype Reiter or immunoblot analyses using the same strain, failed to reveal the presence of the 47 K mol wt antigen in the representative nonpathogenic treponeme. Preabsorption of anti-T. pallidum immune rabbit serum (IRS) with the Reiter organism did not remove anti-T. pallidum antibodies from immune serum that reacted with the 47 K mol wt immunogen or other immunogens of T. pallidum present in the characteristic antigenic profile. Monoclonal antibodies (mAb) directed specifically against the 47 K mol wt immunogen of T. pallidum also failed to react with an analogous 47 K mol wt component in Treponema phagedenis biotype Reiter, further suggesting the unique presence of this antigen in pathogenic treponemes. The presence of the 47 K mol wt surface immunogen in pathogenic treponemes other than T. pallidum subspecies pallidum was also observed (43). Anti-47 K immunogen mAb was nonreactive against rabbit IgG or IgM. mAb directed specifically against the 47 K mol wt immunogen of T. pallidum was examined for strategic functional activities. It was found to be reactive in the microhemagglutination assay for T. pallidum antibodies, the T. pallidum immobilization test, and was found to be capable of significant blockage of attachment of virulent T. pallidum to host cells in tissue culture. Additional significant biological activity for the anti-47 K mol wt immunogen mAb was revealed through results of the in vitro-in vivo neutralization test of Bishop and Miller, in which a 99% or 100% neutralizing activity was demonstrated. The combined data of this study suggest that the 47 K mol wt immunogen of T. pallidum represents an abundant, immunodominant, surface-exposed immunogen possessing potential biological importance in the pathogenesis and immunology of T. pallidum infection. These studies serve to establish the first functionally defined immunogen for T. pallidum, which may represent the major immunogen of the organism.

scholarly journals Lipoproteins Are Responsible for the Pro-Inflammatory Property of Staphylococcus aureus Extracellular Vesicles

2021 ◽  
Vol 22 (13) ◽  
pp. 7099
Author(s):  
Pradeep Kumar Kopparapu ◽  
Meghshree Deshmukh ◽  
Zhicheng Hu ◽  
Majd Mohammad ◽  
Marco Maugeri ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Inflammatory Responses ◽  
Surface Proteins ◽  
Host Cells ◽  
Immune Stimulation ◽  
Domains Of Life ◽  
Major Surface ◽  
Staphylococcal Aureus

Staphylococcal aureus (S. aureus), a Gram-positive bacteria, is known to cause various infections. Extracellular vesicles (EVs) are a heterogeneous array of membranous structures secreted by cells from all three domains of life, i.e., eukaryotes, bacteria, and archaea. Bacterial EVs are implied to be involved in both bacteria–bacteria and bacteria–host interactions during infections. It is still unclear how S. aureus EVs interact with host cells and induce inflammatory responses. In this study, EVs were isolated from S. aureus and mutant strains deficient in either prelipoprotein lipidation (Δlgt) or major surface proteins (ΔsrtAB). Their immunostimulatory capacities were assessed both in vitro and in vivo. We found that S. aureus EVs induced pro-inflammatory responses both in vitro and in vivo. However, this activity was dependent on lipidated lipoproteins (Lpp), since EVs isolated from the Δlgt showed no stimulation. On the other hand, EVs isolated from the ΔsrtAB mutant showed full immune stimulation, indicating the cell wall anchoring of surface proteins did not play a role in immune stimulation. The immune stimulation of S. aureus EVs was mediated mainly by monocytes/macrophages and was TLR2 dependent. In this study, we demonstrated that not only free Lpp but also EV-imbedded Lpp had high pro-inflammatory activity.

scholarly journals Leishmania mexicana Mutants Lacking Glycosylphosphatidylinositol (GPI):Protein Transamidase Provide Insights into the Biosynthesis and Functions of GPI-anchored Proteins

2000 ◽  
Vol 11 (4) ◽  
pp. 1183-1195 ◽  
Author(s):  
James D. Hilley ◽  
Jody L. Zawadzki ◽  
Malcolm J. McConville ◽  
Graham H. Coombs ◽  
Jeremy C. Mottram
Keyword(s):  
Normal Growth ◽  
Surface Proteins ◽  
Host Cells ◽  
Mammalian Host ◽  
Putative Protein ◽  
Major Class ◽  
Major Surface Proteins ◽  
Major Surface

The major surface proteins of the parasitic protozoonLeishmania mexicana are anchored to the plasma membrane by glycosylphosphatidylinositol (GPI) anchors. We have cloned the L. mexicana GPI8 gene that encodes the catalytic component of the GPI:protein transamidase complex that adds GPI anchors to nascent cell surface proteins in the endoplasmic reticulum. Mutants lacking GPI8 (ΔGPI8) do not express detectable levels of GPI-anchored proteins and accumulate two putative protein–anchor precursors. However, the synthesis and cellular levels of other non–protein-linked GPIs, including lipophosphoglycan and a major class of free GPIs, are not affected in the ΔGPI8 mutant. Significantly, the ΔGPI8 mutant displays normal growth in liquid culture, is capable of differentiating into replicating amastigotes within macrophages in vitro, and is infective to mice. These data suggest that GPI-anchored surface proteins are not essential to L. mexicana for its entry into and survival within mammalian host cells in vitro or in vivo and provide further support for the notion that free GPIs are essential for parasite growth.

Immobilization and neutralization of Treponema pallidum attached to cultured mammalian cells

1985 ◽  
Vol 31 (12) ◽  
pp. 1152-1156
Author(s):  
Thomas Fitzgerald
Keyword(s):  
Mammalian Cells ◽  
Neutralizing Antibodies ◽  
Cultured Cells ◽  
Treponema Pallidum ◽  
Immune Serum ◽  
Immune Reactivity ◽  
Complex Environment ◽  
In Vitro Effects

The in vitro effects of antibodies, complement, and (or) macrophages on Treponema pallidum have been previously characterized using relatively simple systems of organisms incubated with the immune components. In vivo, the more complex environment may alter immune reactivity. Experiments were performed to determine whether immobilizing and neutralizing antibodies retained their effectiveness in a more complex environment involving cultured mammalian cells. Two different protocols were used. In protocol A treponemes and normal or immune serum were mixed and added immediately to the cultured cells. In protocol B treponemes were preincubated for 18 h with cultured cells to maximize treponemal attachment; then normal or immune serum was added. With both protocols, attachment of organisms resulted in less effecient immobilization and neutralization. In further experiments, cultured cells were disrupted with Triton X, leaving cytoskeletal remnants on the vessel surface. Identical immobilization and neutralization experiments were performed in the presence of these remnants. In contrast to the findings with viable cultured cells, treponemal attachment to these nonviable remnants did not effect either antibody reaction. Attached organisms were immobilized or neutralized just as efficiently as unattached organisms. Results are discussed in terms of the altered immune reactivity in more complex in vitro environments.

scholarly journals PROPERTIES OF NATIVE PROTEIN-CONTAINING ANTIGENS OF STREPTOCOCCUS PNEUMONIAE

2019 ◽  
Vol 1 (1) ◽  
pp. 22-28
Author(s):  
D. S. Vorobyev ◽  
I. B. Semenova ◽  
Yu. V. Volokh ◽  
E. E. Romanenko ◽  
A. P. Baturo ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Chemical Composition ◽  
Immune Serum ◽  
Rabbit Serum ◽  
Protective Activity ◽  
Native Protein ◽  
In Vivo Experiments ◽  
Sds Electrophoresis

Aim. The study of immunochemical and immunobiological properties of native protein-containing antigens of pneumococcus. Materials and methods. The study was carried out on the strains of the Collective Usage Center «Collection of Mechnikov Res. Inst. for Vaccine and Sera». In the work studied the chemical composition, the molecular weight of the obtained antigens in SDS-electrophoresis and antibody titers in ELISA. Protective activity of protein-containing antigens of pneumococcus was determined in experiments of active protection of mice. Results. Protein-containing antigens of pneumococcus were isolated from S. pneumoniae serotypes 3, 6B, 10A, 14, 19F, 23F and 36. The chemical composition of the preparations contained from 16 to 35% protein. In SDS-electrophoresis in polyacrylamide gel it was established that the molecular weight of protein-containing antigens of pneumococcus ranged from 14 to 116 kDa. Using ELISA shows the cross-activity of native antigens. Virtually all drugs reacted with antimicrobial rabbit serum obtained to serotype 19F (p≤0.05). Serotype serum 14 was less active and only protein-containing pneumococcal antigens obtained from 14 and 19F serotypes (p≤0.05) interacted with it. In the precipitation test according to Ouchterlony it was confirmed that preparations of serotypes 3, 6B, 14, 19F and 36 reacted with rabbit immune serum obtained for S. pneumoniae 19F serotype. In immunoblotting it was found that protein-containing antigens of pneumococcus isolated from serotypes 3, 6B, 10A, 14, 19F and 36 were associated with monoclonal antibodies to pneumococcal protein — pneumolysin. In vivo experiments it was shown that protein-containing antigens of pneumococcus protected animals from intraperitoneal infection of S. pneumoniae in homologous and heterologous systems (p≤0.05). Conclusion. The revealed immunochemical and cross-protective activity of protein-containing antigens of pneumococcus in vitro and in vivo experiments allows to select drugs derived from serotypes 6B, 10A, 19F and 36, as the most promising for further study of the intraspecific protective activity of individual native proteins of pneumococcus. 

scholarly journals ANTIBODY FORMATION AGAINST TREPONEMA PALLIDUM—AGGLUTINATION

1915 ◽  
Vol 21 (6) ◽  
pp. 576-582 ◽  
Author(s):  
Hans Zinsser ◽  
Joseph Gardner Hopkins
Keyword(s):  
Quantitative Difference ◽  
Treponema Pallidum ◽  
Diagnostic Value ◽  
Rabbit Serum ◽  
Normal Rabbit Serum ◽  
Human Beings ◽  
Human Sera ◽  
Normal Human

It has been shown by our experiments that the serum of rabbits treated with emulsions of Treponema pallidum contains agglutinating substances. Normal rabbit serum also possesses agglutinating power for this organism, but, as in the case of normal bacterial agglutinins, to an extent very much inferior to that possessed by the sera of immunized animals. Normal human sera will agglutinate similar pallidum emulsions, as will the sera of certain syphilitic patients with positive Wassermann reactions. Whether or not there is a quantitative difference of diagnostic value between the sera of normal human beings and those of syphilitics remains to be seen. The sera of rabbits immunized with strain A agglutinate Noguchi's strain 9 in dilutions as high as 1 to 500. We regard as the most important result of these experiments the demonstration of definite antibodies in the circulation of animals treated with dead emulsions of Treponema pallidum. Since it is our belief that the agglutinating effect is due to an antibody essentially the same as that which produces bactericidal, precipitating, and opsonic effects, i. e., that there is probably one type of antibody only, we believe that the demonstration of agglutinins establishes the fact that in syphilis as in bacterial diseases the host responds by the formation of antibodies or sensitizers specific for the treponema. Spirocheticidal experiments with these sera, both in vitro and in vivo, are in progress.

Soluble Thrombomodulin Purified from Human Urine Exhibits a Potent Anticoagulant Effect In Vitro and In Vivo

1995 ◽  
Vol 73 (05) ◽  
pp. 805-811 ◽  
Author(s):  
Yasuo Takahashi ◽  
Yoshitaka Hosaka ◽  
Hiromi Niina ◽  
Katsuaki Nagasawa ◽  
Masaaki Naotsuka ◽  
...  
Keyword(s):  
Human Urine ◽  
Specific Activity ◽  
Anticoagulant Activity ◽  
Rabbit Lung ◽  
Soluble Thrombomodulin ◽  
Molecular Weights ◽  
Dose Dependent Fashion ◽  
Dose Dependent

SummaryWe examined the anticoagulant activity of two major molecules of soluble thrombomodulin purified from human urine. The apparent molecular weights of these urinary thrombomodulins (UTMs) were 72,000 and 79,000, respectively. Both UTMs showed more potent cofactor activity for protein C activation [specific activity >5,000 thrombomodulin units (TMU)/mg] than human placental thrombomodulin (2,180 TMU/mg) and rabbit lung thrombomodulin (1,980 TMU/mg). The UTMs prolonged thrombin-induced fibrinogen clotting time (>1 TMU/ml), APTT (>5 TMU/ml), TT (>5 TMU/ml) and PT (>40 TMU/ml) in a dose-dependent fashion. These effects appeared in the concentration range of soluble thrombomodulins present in human plasma and urine. In the rat DIC model induced by thromboplastin, administration of UTMs by infusion (300-3,000 TMU/kg) restored the hematological abnormalities derived from DIC in a dose-dependent fashion. These results demonstrate that UTMs exhibit potent anticoagulant and antithrombotic activities, and could play a physiologically important role in microcirculation.

Toxicity of Heparinoids, with Special Reference to the Precipitation of Fibrinogen

1964 ◽  
Vol 12 (01) ◽  
pp. 232-261 ◽  
Author(s):  
S Sasaki ◽  
T Takemoto ◽  
S Oka
Keyword(s):  
Molecular Weight ◽  
Dextran Sulfate ◽  
Anticoagulant Activity ◽  
Molecular Structures ◽  
Severe Reaction ◽  
Clinical Use ◽  
Molecular Weights ◽  
Close Relationship

SummaryTo demonstrate whether the intravascular precipitation of fibrinogen is responsible for the toxicity of heparinoid, the relation between the toxicity of heparinoid in vivo and the precipitation of fibrinogen in vitro was investigated, using dextran sulfate of various molecular weights and various heparinoids.1. There are close relationships between the molecular weight of dextran sulfate, its toxicity, and the quantity of fibrinogen precipitated.2. The close relationship between the toxicity and the precipitation of fibrinogen found for dextran sulfate holds good for other heparinoids regardless of their molecular structures.3. Histological findings suggest strongly that the pathological changes produced with dextran sulfate are caused primarily by the intravascular precipitates with occlusion of the capillaries.From these facts, it is concluded that the precipitates of fibrinogen with heparinoid may be the cause or at least the major cause of the toxicity of heparinoid.4. The most suitable molecular weight of dextran sulfate for clinical use was found to be 5,300 ~ 6,700, from the maximum value of the product (LD50 · Anticoagulant activity). This product (LD50 · Anticoagulant activity) can be employed generally to assess the comparative merits of various heparinoids.5. Clinical use of the dextran sulfate prepared on this basis gave satisfactory results. No severe reaction was observed. However, two delayed reactions, alopecia and thrombocytopenia, were observed. These two reactions seem to come from the cause other than intravascular precipitation.

ACIS, A Novel KepTide™, Binds to ACE-2 Receptor and Inhibits the Infection of SARS-CoV2 Virus in vitro in Primate Kidney Cells: Therapeutic Implications for COVID-19 (Preprint)

2020 ◽  
Author(s):  
Avik Sotira Scientific
Keyword(s):  
Social Life ◽  
Plaque Assay ◽  
Host Cells ◽  
Surface Glycoprotein ◽  
Crystallographic Structure ◽  
Therapeutic Implications ◽  
Biochemical Assays ◽  
Targeted Medicine

UNSTRUCTURED Coronavirus disease 2019 (COVID-19) is a severe acute respiratory syndrome (SARS) caused by a virus known as SARS-Coronavirus 2 (SARS-CoV2). Without a targeted-medicine, this disease has been causing a massive humanitarian crisis not only in terms of mortality, but also imposing a lasting damage to social life and economic progress of humankind. Therefore, an immediate therapeutic strategy needs to be intervened to mitigate this global crisis. Here, we report a novel KepTide™ (Knock-End Peptide) therapy that nullifies SARS-CoV2 infection. SARS-CoV2 employs its surface glycoprotein “spike” (S-glycoprotein) to interact with angiotensin converting enzyme-2 (ACE-2) receptor for its infection in host cells. Based on our in-silico-based homology modeling study validated with a recent X-ray crystallographic structure (PDB ID:6M0J), we have identified that a conserved motif of S-glycoprotein that intimately engages multiple hydrogen-bond (H-bond) interactions with ACE-2 enzyme. Accordingly, we designed a peptide, termed as ACIS (ACE-2 Inhibitory motif of Spike), that displayed significant affinity towards ACE-2 enzyme as confirmed by biochemical assays such as BLItz and fluorescence polarization assays. Interestingly, more than one biochemical modifications were adopted in ACIS in order to enhance the inhibitory action of ACIS and hence called as KEpTide™. Consequently, a monolayer invasion assay, plaque assay and dual immunofluorescence analysis further revealed that KEpTide™ efficiently mitigated the infection of SARS-CoV2 in vitro in VERO E6 cells. Finally, evaluating the relative abundance of ACIS in lungs and the potential side-effects in vivo in mice, our current study discovers a novel KepTide™ therapy that is safe, stable, and robust to attenuate the infection of SARS-CoV2 virus if administered intranasally. INTERNATIONAL REGISTERED REPORT RR2-https://doi.org/10.1101/2020.10.13.337584

scholarly journals Effective decellularisation of human saphenous veins for biocompatible arterial tissue engineering applications: Bench optimisation and feasibility in vivo testing

2021 ◽  
Vol 12 ◽  
pp. 204173142098752
Author(s):  
Nadiah S Sulaiman ◽  
Andrew R Bond ◽  
Vito D Bruno ◽  
John Joseph ◽  
Jason L Johnson ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Mechanical Strength ◽  
Vascular Tissue ◽  
Sodium Dodecyl ◽  
Host Cells ◽  
Replacement Model ◽  
In Vivo Testing ◽  
Vascular Scaffolds

Human saphenous vein (hSV) and synthetic grafts are commonly used conduits in vascular grafting, despite high failure rates. Decellularising hSVs (D-hSVs) to produce vascular scaffolds might be an effective alternative. We assessed the effectiveness of a detergent-based method using 0% to 1% sodium dodecyl sulphate (SDS) to decellularise hSV. Decellularisation effectiveness was measured in vitro by nuclear counting, DNA content, residual cell viability, extracellular matrix integrity and mechanical strength. Cytotoxicity was assessed on human and porcine cells. The most effective SDS concentration was used to prepare D-hSV grafts that underwent preliminary in vivo testing using a porcine carotid artery replacement model. Effective decellularisation was achieved with 0.01% SDS, and D-hSVs were biocompatible after seeding. In vivo xeno-transplantation confirmed excellent mechanical strength and biocompatibility with recruitment of host cells without mechanical failure, and a 50% patency rate at 4-weeks. We have developed a simple biocompatible methodology to effectively decellularise hSVs. This could enhance vascular tissue engineering toward future clinical applications.

scholarly journals P074 In vitro and in vivo efficacy of linezolid on Treponema pallidum, the syphilis agent

2021 ◽  
Author(s):  
L Giacani ◽  
A Haynes ◽  
M Vall Mayans ◽  
M Ubals Cazorla ◽  
C Nieto ◽  
...  
Keyword(s):  
Treponema Pallidum ◽  
In Vivo Efficacy
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