The inhibitory effect of cromolyn sodium and ketotifen on Toxoplasma gondii entrance into host cells in vitro and in vivo

Toxoplasma gondii secretes a number of virulence-related effector proteins, such as the rhoptry protein 18 (ROP18). To further broaden our understanding of the molecular functions of ROP18, we examined the transcriptional response of human embryonic kidney cells (HEK293T) to ROP18 of type I T. gondii RH strain. Using RNA-sequencing, we compared the transcriptome of ROP18-expressing HEK293T cells to control HEK293T cells. Our analysis revealed that ROP18 altered the expression of 750 genes (467 upregulated genes and 283 downregulated genes) in HEK293T cells. Gene ontology (GO) and pathway enrichment analyses showed that differentially expressed genes (DEGs) were significantly enriched in extracellular matrix– and immune–related GO terms and pathways. KEGG pathway enrichment analysis revealed that DEGs were involved in several disease-related pathways, such as nervous system diseases and eye disease. ROP18 significantly increased the alternative splicing pattern “retained intron” and altered the expression of 144 transcription factors (TFs). These results provide new insight into how ROP18 may influence biological processes in the host cells via altering the expression of genes, TFs, and pathways. More in vitro and in vivo studies are required to substantiate these findings.

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The ketolide antibiotics HMR 3647 and HMR 3004 are active against Toxoplasma gondii in vitro and in murine models of infection.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.41.10.2137 ◽

1997 ◽

Vol 41 (10) ◽

pp. 2137-2140 ◽

Cited By ~ 30

Author(s):

F G Araujo ◽

A A Khan ◽

T L Slifer ◽

A Bryskier ◽

J S Remington

Keyword(s):

Toxoplasma Gondii ◽

Protozoan Parasite ◽

Host Cells ◽

Resistant Bacteria ◽

New Class ◽

Human Foreskin Fibroblasts ◽

Significant Toxicity ◽

Different Strains

Ketolides are a new class of macrolide antibiotics that have been shown to be active against a variety of bacteria including macrolide-resistant bacteria and mycobacteria. We examined two ketolides, HMR 3647 and HMR 3004, for their in vitro and in vivo activities against the protozoan parasite Toxoplasma gondii. In vitro, both ketolides at concentrations as low as 0.05 microg/ml markedly inhibited replication of tachyzoites of the RH strain within human foreskin fibroblasts. HMR 3004 demonstrated some toxicity for host cells after they were exposed to 5 microg of the drug per ml for 72 h. In contrast, HMR 3647 did not show any significant toxicity even at concentrations as high as 25 microg/ml. In vivo, both ketolides provided remarkable protection against death in mice lethally infected intraperitoneally with tachyzoites of the RH strain or orally with tissue cysts of the C56 strain of T. gondii. A dosage of 100 mg of HMR 3647 per kg of body weight per day administered for 10 days protected 50% of mice infected with tachyzoites. The same dosage of HMR 3004 protected 100% of the mice. In mice infected with cysts, a dosage of 30 mg of HMR 3647 per kg per day protected 100% of the mice, whereas a dosage of 40 mg of HMR 3004 per kg per day protected 75% of the mice. These results demonstrate that HMR 3647 and HMR 3004 possess excellent activities against two different strains of T. gondii and may be useful for the treatment of toxoplasmosis in humans.

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Role of F1C Fimbriae, Flagella, and Secreted Bacterial Components in the Inhibitory Effect of Probiotic Escherichia coli Nissle 1917 on Atypical Enteropathogenic E. coli Infection

Infection and Immunity ◽

10.1128/iai.01431-13 ◽

2014 ◽

Vol 82 (5) ◽

pp. 1801-1812 ◽

Cited By ~ 23

Author(s):

Sylvia Kleta ◽

Marcel Nordhoff ◽

Karsten Tedin ◽

Lothar H. Wieler ◽

Rafal Kolenda ◽

...

Keyword(s):

Escherichia Coli ◽

Molecular Mechanisms ◽

Inhibitory Effect ◽

Host Cells ◽

Single Infection ◽

Content Type ◽

E Coli ◽

Initial Adhesion

ABSTRACTEnteropathogenicEscherichia coli(EPEC) is recognized as an important intestinal pathogen that frequently causes acute and persistent diarrhea in humans and animals. The use of probiotic bacteria to prevent diarrhea is gaining increasing interest. The probioticE. colistrain Nissle 1917 (EcN) is known to be effective in the treatment of several gastrointestinal disorders. While bothin vitroandin vivostudies have described strong inhibitory effects of EcN on enteropathogenic bacteria, including pathogenicE. coli, the underlying molecular mechanisms remain largely unknown. In this study, we examined the inhibitory effect of EcN on infections of porcine intestinal epithelial cells with atypical enteropathogenicE. coli(aEPEC) with respect to single infection steps, including adhesion, microcolony formation, and the attaching and effacing phenotype. We show that EcN drastically reduced the infection efficiencies of aEPEC by inhibiting bacterial adhesion and growth of microcolonies, but not the attaching and effacing of adherent bacteria. The inhibitory effect correlated with EcN adhesion capacities and was predominantly mediated by F1C fimbriae, but also by H1 flagella, which served as bridges between EcN cells. Furthermore, EcN seemed to interfere with the initial adhesion of aEPEC to host cells by secretion of inhibitory components. These components do not appear to be specific to EcN, but we propose that the strong adhesion capacities enable EcN to secrete sufficient local concentrations of the inhibitory factors. The results of this study are consistent with a mode of action whereby EcN inhibits secretion of virulence-associated proteins of EPEC, but not their expression.

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Actin depolymerizing factor controls actin turnover and gliding motility in Toxoplasma gondii

Molecular Biology of the Cell ◽

10.1091/mbc.e10-12-0939 ◽

2011 ◽

Vol 22 (8) ◽

pp. 1290-1299 ◽

Cited By ~ 42

Author(s):

Simren Mehta ◽

L. David Sibley

Keyword(s):

Toxoplasma Gondii ◽

Actin Filaments ◽

Gliding Motility ◽

Conditional Knockout ◽

Host Cells ◽

Actin Binding ◽

Actin Depolymerizing Factor

Apicomplexan parasites rely on actin-based gliding motility to move across the substratum, cross biological barriers, and invade their host cells. Gliding motility depends on polymerization of parasite actin filaments, yet ∼98% of actin is nonfilamentous in resting parasites. Previous studies suggest that the lack of actin filaments in the parasite is due to inherent instability, leaving uncertain the role of actin-binding proteins in controlling dynamics. We have previously shown that the single allele of Toxoplasma gondii actin depolymerizing factor (TgADF) has strong actin monomer–sequestering and weak filament-severing activities in vitro. Here we used a conditional knockout strategy to investigate the role of TgADF in vivo. Suppression of TgADF led to accumulation of actin-rich filaments that were detected by immunofluorescence and electron microscopy. Parasites deficient in TgADF showed reduced speed of motility, increased aberrant patterns of motion, and inhibition of sustained helical gliding. Lack of TgADF also led to severe defects in entry and egress from host cells, thus blocking infection in vitro. These studies establish that the absence of stable actin structures in the parasite are not simply the result of intrinsic instability, but that TgADF is required for the rapid turnover of parasite actin filaments, gliding motility, and cell invasion.

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p47 GTPases Regulate Toxoplasma gondii Survival in Activated Macrophages

Infection and Immunity ◽

10.1128/iai.73.6.3278-3286.2005 ◽

2005 ◽

Vol 73 (6) ◽

pp. 3278-3286 ◽

Cited By ~ 99

Author(s):

Barbara A. Butcher ◽

Robert I. Greene ◽

Stanley C. Henry ◽

Kimberly L. Annecharico ◽

J. Brice Weinberg ◽

...

Keyword(s):

Toxoplasma Gondii ◽

Acute Infection ◽

Host Cells ◽

Intracellular Growth ◽

Latex Beads ◽

Cellular Factors ◽

Ifn Γ ◽

Normal Inhibition

ABSTRACT The cytokine gamma interferon (IFN-γ) is critical for resistance to Toxoplasma gondii. IFN-γ strongly activates macrophages and nonphagocytic host cells to limit intracellular growth of T. gondii; however, the cellular factors that are required for this effect are largely unknown. We have shown previously that IGTP and LRG-47, members of the IFN-γ-regulated family of p47 GTPases, are required for resistance to acute T. gondii infections in vivo. In contrast, IRG-47, another member of this family, is not required. In the present work, we addressed whether these GTPases are required for IFN-γ-induced suppression of T. gondii growth in macrophages in vitro. Bone marrow macrophages that lacked IGTP or LRG-47 displayed greatly attenuated IFN-γ-induced inhibition of T. gondii growth, while macrophages that lacked IRG-47 displayed normal inhibition. Thus, the ability of the p47 GTPases to limit acute infection in vivo correlated with their ability to suppress intracellular growth in macrophages in vitro. Using confocal microscopy and sucrose density fractionation, we demonstrated that IGTP largely colocalizes with endoplasmic reticulum markers, while LRG-47 was mainly restricted to the Golgi. Although both IGTP and LRG-47 localized to vacuoles containing latex beads, neither protein localized to vacuoles containing live T. gondii. These results suggest that IGTP and LRG-47 are able to regulate host resistance to acute T. gondii infections through their ability to inhibit parasite growth within the macrophage.

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Anti-Toxoplasma gondii Effects of a Novel Spider Peptide XYP1 In Vitro and In Vivo

Biomedicines ◽

10.3390/biomedicines9080934 ◽

2021 ◽

Vol 9 (8) ◽

pp. 934

Author(s):

Yuan Liu ◽

Yaqin Tang ◽

Xing Tang ◽

Mengqi Wu ◽

Shengjie Hou ◽

...

Keyword(s):

Electron Microscopy ◽

Toxoplasma Gondii ◽

Venom Gland ◽

Host Cells ◽

Drug Candidate ◽

Membrane Perforation ◽

Associated Proteins ◽

Low Cytotoxicity

Toxoplasmosis, caused by an obligate intracellular parasite Toxoplasma gondii, is one of the most prevalent zoonoses worldwide. Treatments for this disease by traditional drugs have shown numerous side effects, thus effective alternative anti-Toxoplasma strategies or drugs are urgently needed. In this study, a novel spider peptide, XYP1, was identified from the cDNA library of the venom gland of the spider Lycosa coelestis. Our results showed that XYP1 has potent anti-Toxoplasma activity in vitro and in vivo. Specifically, treatment with XYP1 significantly inhibited the viability, invasion and proliferation of tachyzoites with low cytotoxicity (IC50 = 38.79 μΜ) on human host cells, and increased the survival rate of mice acutely infected with T. gondii. Next, scanning electron microscopy, transmission electron microscopy and RNA sequencing were employed to further explore the functional mechanism of XYP1, and the results indicated that XYP1 causes membrane perforation, swelling and disruption of tachyzoites, which could be closely associated with differential expression of several membrane-associated proteins including HSP29. In conclusion, XYP1 may be a promising new drug candidate for the treatment of toxoplasmosis.

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Rifapentine is active in vitro and in vivo against Toxoplasma gondii.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.40.6.1335 ◽

1996 ◽

Vol 40 (6) ◽

pp. 1335-1337 ◽

Cited By ~ 24

Author(s):

F G Araujo ◽

A A Khan ◽

J S Remington

Keyword(s):

Toxoplasma Gondii ◽

Protozoan Parasite ◽

Host Cells ◽

Intracellular Replication ◽

Degree Of Protection

Rifapentine, a derivative of rifamycin, was examined for its in vitro and in vivo activities against the protozoan parasite Toxoplasma gondii. The drug inhibited the intracellular replication of parasites and was not cytotoxic for the host cells at inhibitory concentrations. Mice infected either intraperitoneally with tachyzoites of the RH strain or orally with tissue cysts of the C56 strain were protected against death by treatment with rifapentine. The degree of protection was similar to that induced by atovaquone and apparently higher than that induced by rifabutin. Rifapentine may be a useful drug for the treatment of toxoplasmosis in immunocompromised individuals.

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AMA1-Deficient Toxoplasma gondii Parasites Transiently Colonize Mice and Trigger an Innate Immune Response That Leads to Long-Lasting Protective Immunity

Infection and Immunity ◽

10.1128/iai.02606-14 ◽

2015 ◽

Vol 83 (6) ◽

pp. 2475-2486 ◽

Cited By ~ 14

Author(s):

Vanessa Lagal ◽

Márcia Dinis ◽

Dominique Cannella ◽

Daniel Bargieri ◽

Virginie Gonzalez ◽

...

Keyword(s):

Toxoplasma Gondii ◽

Distinct Type ◽

Genetically Engineered ◽

Host Cells ◽

Type I ◽

Content Type ◽

Apical Membrane Antigen 1 ◽

Immunocompromised Mice

The apical membrane antigen 1 (AMA1) protein was believed to be essential for the perpetuation of two Apicomplexa parasite genera,PlasmodiumandToxoplasma, until we genetically engineered viable parasites lackingAMA1. The reduction in invasiveness of theToxoplasma gondiiRH-AMA1 knockout (RH-AMA1KO) tachyzoite population,in vitro, raised key questions about the outcome associated with these tachyzoites once inoculated in the peritoneal cavity of mice. In this study, we used AMNIS technology to simultaneously quantify and image the parasitic process driven by AMA1KOtachyzoites. We report their ability to colonize and multiply in mesothelial cells and in both resident and recruited leukocytes. While the RH-AMA1KOpopulation amplification is rapidly lethal in immunocompromised mice, it is controlled in immunocompetent hosts, where immune cells in combination sense parasites and secrete proinflammatory cytokines. This innate response further leads to a long-lasting status immunoprotective against a secondary challenge by high inocula of the homologous type I or a distinct type IIT. gondiigenotypes. While AMA1 is definitively not an essential protein for tachyzoite entry and multiplication in host cells, it clearly assists the expansion of parasite populationin vivo.

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In vitro action of antiparasitic drugs, especially artesunate, against Toxoplasma gondii

Revista da Sociedade Brasileira de Medicina Tropical ◽

10.1590/s0037-86822012000400014 ◽

2012 ◽

Vol 45 (4) ◽

pp. 485-490 ◽

Cited By ~ 8

Author(s):

Thaís Cobellis Gomes ◽

Heitor Franco de Andrade Júnior ◽

Susana Angélica Zevallos Lescano ◽

Vicente Amato-Neto

Keyword(s):

Toxoplasma Gondii ◽

Inhibitory Concentration ◽

Inhibitory Effect ◽

Congenital Disease ◽

Nonlinear Regression Analysis ◽

Antiparasitic Drugs ◽

Effective Drugs ◽

Disposable Plastic

INTRODUCTION: Toxoplasmosis is usually a benign infection, except in the event of ocular, central nervous system (CNS), or congenital disease and particularly when the patient is immunocompromised. Treatment consists of drugs that frequently cause adverse effects; thus, newer, more effective drugs are needed. In this study, the possible activity of artesunate, a drug successfully being used for the treatment of malaria, on Toxoplasma gondii growth in cell culture is evaluated and compared with the action of drugs that are already being used against this parasite. METHODS: LLC-MK2 cells were cultivated in RPMI medium, kept in disposable plastic bottles, and incubated at 36ºC with 5% CO2. Tachyzoites of the RH strain were used. The following drugs were tested: artesunate, cotrimoxazole, pentamidine, pyrimethamine, quinine, and trimethoprim. The effects of these drugs on tachyzoites and LLC-MK2 cells were analyzed using nonlinear regression analysis with Prism 3.0 software. RESULTS: Artesunate showed a mean tachyzoite inhibitory concentration (IC50) of 0.075µM and an LLC MK2 toxicity of 2.003µM. Pyrimethamine was effective at an IC50 of 0.482µM and a toxicity of 11.178µM. Trimethoprim alone was effective against the in vitro parasite. Cotrimoxazole also was effective against the parasite but at higher concentrations than those observed for artesunate and pyrimethamine. Pentamidine and quinine had no inhibitory effect over tachyzoites. CONCLUSIONS: Artesunate is proven in vitro to be a useful alternative for the treatment of toxoplasmosis, implying a subsequent in vivo effect and suggesting the mechanism of this drug against the parasite.

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Toxoplasma gondii co-opts the unfolded protein response to enhance migration and dissemination of infected host cells

10.1101/2020.04.14.042069 ◽

2020 ◽

Author(s):

Leonardo Augusto ◽

Jennifer Martynowicz ◽

Parth H. Amin ◽

Nada S. Alakhras ◽

Mark H. Kaplan ◽

...

Keyword(s):

Toxoplasma Gondii ◽

Host Cell ◽

The Body ◽

Host Cells ◽

Migratory Activity ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

AbstractToxoplasma gondii is an intracellular parasite that reconfigures its host cell to promote pathogenesis. One consequence of Toxoplasma parasitism is increased migratory activity of host cells, which facilitates dissemination. Here we show that Toxoplasma triggers the unfolded protein response (UPR) in host cells through calcium release from the endoplasmic reticulum (ER). We further found that host IRE1, an ER stress sensor protein activated during Toxoplasma infection, also plays a noncanonical role in actin remodeling by binding filamin A in infected cells. By inducing cytoskeletal remodeling via IRE1 oligomerization in host cells, Toxoplasma enhances host cell migration in vitro and dissemination of the parasite to host organs in vivo. Our study identifies novel mechanisms used by Toxoplasma to induce dissemination of infected cells, providing new insights into strategies for treatment of toxoplasmosis.ImportanceCells that are infected with the parasite Toxoplasma gondii exhibit heightened migratory activity, which facilitates dissemination of the infection throughout the body. In this study, we identify a new mechanism used by Toxoplasma to hijack its host cell and increase its mobility. We further show that the ability of Toxoplasma to increase host cell migration does not involve the enzymatic activity of IRE1, but rather IRE1 engagement with actin cytoskeletal remodeling. Depletion of IRE1 from infected host cells reduces their migration in vitro and significantly hinders dissemination of Toxoplasma in vivo. Our findings reveal a new mechanism underlying host-pathogen interactions, demonstrating how host cells are co-opted to spread a persistent infection around the body.

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