scholarly journals Migratory Activation of Primary Cortical Microglia upon Infection with Toxoplasma gondii

2011 ◽  
Vol 79 (8) ◽  
pp. 3046-3052 ◽  
Author(s):  
Isabel Dellacasa-Lindberg ◽  
Jonas M. Fuks ◽  
Romanico B. G. Arrighi ◽  
Henrik Lambert ◽  
Robert P. A. Wallin ◽  
...  
Keyword(s):  
T Cells ◽  
Toxoplasma Gondii ◽  
Mhc Class Ii ◽  
Glial Cells ◽  
Costimulatory Molecule ◽  
Toxoplasmic Encephalitis ◽  
Immune Functions ◽  
Content Type

ABSTRACTDisseminated toxoplasmosis in the central nervous system (CNS) is often accompanied by a lethal outcome. Studies with murine models of infection have focused on the role of systemic immunity in control of toxoplasmic encephalitis, while knowledge remains limited on the contributions of resident cells with immune functions in the CNS. In this study, the role of glial cells was addressed in the setting of recrudescentToxoplasmainfection in mice. Activated astrocytes and microglia were observed in the close vicinity of foci with replicating parasitesin situin the brain parenchyma.Toxoplasma gondiitachyzoites were allowed to infect primary microglia and astrocytesin vitro. Microglia were permissive to parasite replication, and infected microglia readily transmigrated across transwell membranes and cell monolayers. Thus, infected microglia, but not astrocytes, exhibited a hypermotility phenotype reminiscent of that recently described for infected dendritic cells. In contrast to gamma interferon-activated microglia,Toxoplasma-infected microglia did not upregulate major histocompatibility complex (MHC) class II molecules and the costimulatory molecule CD86. YetToxoplasma-infected microglia and astrocytes exhibited increased sensitivity to T cell-mediated killing, leading to rapid parasite transfer to effector T cellsin vitro. We hypothesize that glial cells and T cells, besides their role in triggering antiparasite immunity, may also act as “Trojan horses,” paradoxically facilitating dissemination ofToxoplasmawithin the CNS. To our knowledge, this constitutes the first report of migratory activation of a resident CNS cell by an intracellular parasite.

scholarly journals Role of Toxoplasma gondii Chloroquine Resistance Transporter in Bradyzoite Viability and Digestive Vacuole Maintenance

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Geetha Kannan ◽  
Manlio Di Cristina ◽  
Aric J. Schultz ◽  
My-Hang Huynh ◽  
Fengrong Wang ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Chronic Infection ◽  
Chloroquine Resistance ◽  
Congenital Defects ◽  
Functional Link ◽  
Content Type ◽  
Chloroquine Resistance Transporter

ABSTRACT Toxoplasma gondii is a ubiquitous pathogen that can cause encephalitis, congenital defects, and ocular disease. T. gondii has also been implicated as a risk factor for mental illness in humans. The parasite persists in the brain as slow-growing bradyzoites contained within intracellular cysts. No treatments exist to eliminate this form of parasite. Although proteolytic degradation within the parasite lysosome-like vacuolar compartment (VAC) is critical for bradyzoite viability, whether other aspects of the VAC are important for parasite persistence remains unknown. An ortholog of Plasmodium falciparum chloroquine resistance transporter (CRT), TgCRT, has previously been identified in T. gondii. To interrogate the function of TgCRT in chronic-stage bradyzoites and its role in persistence, we knocked out TgCRT in a cystogenic strain and assessed VAC size, VAC digestion of host-derived proteins and parasite autophagosomes, and the viability of in vitro and in vivo bradyzoites. We found that whereas parasites deficient in TgCRT exhibit normal digestion within the VAC, they display a markedly distended VAC and their viability is compromised both in vitro and in vivo. Interestingly, impairing VAC proteolysis in TgCRT-deficient bradyzoites restored VAC size, consistent with a role for TgCRT as a transporter of products of digestion from the VAC. In conjunction with earlier studies, our current findings suggest a functional link between TgCRT and VAC proteolysis. This study provides further evidence of a crucial role for the VAC in bradyzoite persistence and a new potential VAC target to abate chronic Toxoplasma infection. IMPORTANCE Individuals chronically infected with the intracellular parasite Toxoplasma gondii are at risk of experiencing reactivated disease that can result in progressive loss of vision. No effective treatments exist for chronic toxoplasmosis due in part to a poor understanding of the biology underlying chronic infection and a lack of well-validated potential targets. We show here that a T. gondii transporter is functionally linked to protein digestion within the parasite lysosome-like organelle and that this transporter is necessary to sustain chronic infection in culture and in experimentally infected mice. Ablating the transporter results in severe bloating of the lysosome-like organelle. Together with earlier work, this study suggests the parasite’s lysosome-like organelle is vital for parasite survival, thus rendering it a potential target for diminishing infection and reducing the risk of reactivated disease.

scholarly journals Significant Reduction of Brain Cysts Caused by Toxoplasma gondii after Treatment with Spiramycin Coadministered with Metronidazole in a Mouse Model of Chronic Toxoplasmosis

2012 ◽  
Vol 56 (4) ◽  
pp. 1762-1768 ◽  
Author(s):  
Wai Kit Chew ◽  
Ignacio Segarra ◽  
Stephen Ambu ◽  
Joon Wah Mak
Keyword(s):  
Toxoplasma Gondii ◽  
Vero Cells ◽  
Toxoplasmic Encephalitis ◽  
Combination Group ◽  
Control Group ◽  
Time Curve ◽  
Content Type ◽  
Brain Cysts ◽  
The Brain

ABSTRACTToxoplasma gondiiis a parasite that generates latent cysts in the brain; reactivation of these cysts may lead to fatal toxoplasmic encephalitis, for which treatment remains unsuccessful. We assessed spiramycin pharmacokinetics coadministered with metronidazole, the eradication of brain cysts and thein vitroreactivation. Male BALB/c mice were fed 1,000 tachyzoites orally to develop chronic toxoplasmosis. Four weeks later, infected mice underwent different treatments: (i) infected untreated mice (n= 9), which received vehicle only; (ii) a spiramycin-only group (n= 9), 400 mg/kg daily for 7 days; (iii) a metronidazole-only group (n= 9), 500 mg/kg daily for 7 days; and (iv) a combination group (n= 9), which received both spiramycin (400 mg/kg) and metronidazole (500 mg/kg) daily for 7 days. An uninfected control group (n= 10) was administered vehicle only. After treatment, the brain cysts were counted, brain homogenates were cultured in confluent Vero cells, and cysts and tachyzoites were counted after 1 week. Separately, pharmacokinetic profiles (plasma and brain) were assessed after a single dose of spiramycin (400 mg/kg), metronidazole (500 mg/kg), or both. Metronidazole treatment increased the brain spiramycin area under the concentration-time curve from 0 h to ∞ (AUC0–∞) by 67% without affecting its plasma disposition. Metronidazole plasma and brain AUC0–∞values were reduced 9 and 62%, respectively, after spiramycin coadministration. Enhanced spiramycin brain exposure after coadministration reduced brain cysts 15-fold (79 ± 23 for the combination treatment versus 1,198 ± 153 for the untreated control group [P< 0.05]) and 10-fold versus the spiramycin-only group (768 ± 125). Metronidazole alone showed no effect (1,028 ± 149). Tachyzoites were absent in the brain. Spiramycin reducedin vitroreactivation. Metronidazole increased spiramycin brain penetration, causing a significant reduction ofT. gondiibrain cysts, with potential clinical translatability for chronic toxoplasmosis treatment.

scholarly journals VCAM-1/α4β1 Integrin Interaction Is Crucial for Prompt Recruitment of Immune T Cells into the Brain during the Early Stage of Reactivation of Chronic Infection with Toxoplasma gondii To Prevent Toxoplasmic Encephalitis

2014 ◽  
Vol 82 (7) ◽  
pp. 2826-2839 ◽  
Author(s):  
Qila Sa ◽  
Eri Ochiai ◽  
Tomoko Sengoku ◽  
Melinda E. Wilson ◽  
Morgan Brogli ◽  
...  
Keyword(s):  
T Cells ◽  
Toxoplasma Gondii ◽  
Chronic Infection ◽  
Early Stage ◽  
Toxoplasmic Encephalitis ◽  
Mrna Levels ◽  
Wild Type ◽  
Content Type ◽  
Ifn Γ ◽  
The Brain

ABSTRACTReactivation of chronic infection withToxoplasma gondiican cause life-threatening toxoplasmic encephalitis in immunocompromised individuals. We examined the role of VCAM-1/α4β1 integrin interaction in T cell recruitment to prevent reactivation of the infection in the brain. SCID mice were infected and treated with sulfadiazine to establish a chronic infection. VCAM-1 and ICAM-1 were the endothelial adhesion molecules detected on cerebral vessels of the infected SCID and wild-type animals. Immune T cells from infected wild-type mice were treated with anti-α4 integrin or control antibodies and transferred into infected SCID or nude mice, and the animals received the same antibody every other day. Three days later, sulfadiazine was discontinued to initiate reactivation of infection. Expression of mRNAs for CD3δ, CD4, CD8β, gamma interferon (IFN-γ), and inducible nitric oxide synthase (NOS2) (an effector molecule to inhibitT. gondiigrowth) and the numbers of CD4+and CD8+T cells in the brain were significantly less in mice treated with anti-α4 integrin antibody than in those treated with control antibody at 3 days after sulfadiazine discontinuation. At 6 days after sulfadiazine discontinuation, cerebral tachyzoite-specific SAG1 mRNA levels and numbers of inflammatory foci associated with tachyzoites were markedly greater in anti-α4 integrin antibody-treated than in control antibody-treated animals, even though IFN-γ and NOS2 mRNA levels were higher in the former than in the latter. These results indicate that VCAM-1/α4β1 integrin interaction is crucial for prompt recruitment of immune T cells and induction of IFN-γ-mediated protective immune responses during the early stage of reactivation of chronicT. gondiiinfection to control tachyzoite growth.

scholarly journals Nonpathogenic Colonization with Chlamydia in the Gastrointestinal Tract as Oral Vaccination for Inducing Transmucosal Protection

2017 ◽  
Vol 86 (2) ◽  
Author(s):  
Luying Wang ◽  
Cuiming Zhu ◽  
Tianyuan Zhang ◽  
Qi Tian ◽  
Nu Zhang ◽  
...  
Keyword(s):  
T Cells ◽  
Gastrointestinal Tract ◽  
Antigen Presentation ◽  
Mhc Class Ii ◽  
Genital Tract ◽  
Genital Tract Infection ◽  
Content Type ◽  
Chlamydia Muridarum ◽  
Class Ii Antigen

ABSTRACTChlamydiahas been detected in the gastrointestinal tracts of humans and animals. We now report that gastrointestinalChlamydia muridarumis able to induce robust transmucosal protection in mice.C. muridarumcolonization in the gastrointestinal tract correlated with both a shortened course ofC. muridarumgenital tract infection and stronger protection against subsequent genital tract challenge infection. Mice preinoculated intragastrically withC. muridarumbecame highly resistant to subsequentC. muridaruminfection in the genital tract, resulting in prevention of pathology in the upper genital tract. The transmucosal protection in the genital tract was rapidly induced, durable, and dependent on major histocompatibility complex (MHC) class II antigen presentation but not MHC class I antigen presentation. Although a deficiency in CD4+T cells only partially reduced the transmucosal protection, depletion of CD4+T cells from B cell-deficient mice completely abolished the protection, suggesting a synergistic role of both CD4+T and B cells in the gastrointestinalC. muridarum-induced transmucosal immunity. However, the same protective immunity did not significantly affectC. muridarumcolonization in the gastrointestinal tract. The long-lasting colonization withC. muridarumwas restricted to the gastrointestinal tract and was nonpathogenic to either gastrointestinal or extragastrointestinal tissues. Furthermore, gastrointestinalC. muridarumdid not alter the gut microbiota or the development of gut mucosal resident memory T cell responses to a nonchlamydial infection. Thus,Chlamydiamay be developed into a safe and orally deliverable replicating vaccine for inducing transmucosal protection.

scholarly journals Transcriptional Profiling Suggests T Cells Cluster around Neurons Injected with Toxoplasma gondii Proteins

mSphere ◽  
2020 ◽  
Vol 5 (5) ◽  
Author(s):  
Emily F. Merritt ◽  
Hannah J. Johnson ◽  
Zhee Sheen Wong ◽  
Adam S. Buntzman ◽  
Austin C. Conklin ◽  
...  
Keyword(s):  
T Cells ◽  
Toxoplasma Gondii ◽  
Immune Cells ◽  
Drug Targets ◽  
Transcriptional Profiling ◽  
Effector Proteins ◽  
Content Type ◽  
Symptomatic Disease

ABSTRACT Toxoplasma gondii’s tropism for and persistence in the central nervous system (CNS) underlies the symptomatic disease that T. gondii causes in humans. Our recent work has shown that neurons are the primary CNS cell with which Toxoplasma interacts and which it infects in vivo. This predilection for neurons suggests that T. gondii’s persistence in the CNS depends specifically upon parasite manipulation of the host neurons. Yet, most work on T. gondii-host cell interactions has been done in vitro and in nonneuronal cells. We address this gap by utilizing our T. gondii-Cre system that allows permanent marking and tracking of neurons injected with parasite effector proteins in vivo. Using laser capture microdissection (LCM) and RNA sequencing using RNA-seq, we isolated and transcriptionally profiled T. gondii-injected neurons (TINs), Bystander neurons (nearby non-T. gondii-injected neurons), and neurons from uninfected mice (controls). These profiles show that TIN transcriptomes significantly differ from the transcriptomes of Bystander and control neurons and that much of this difference is driven by increased levels of transcripts from immune cells, especially CD8+ T cells and monocytes. These data suggest that when we used LCM to isolate neurons from infected mice, we also picked up fragments of CD8+ T cells and monocytes clustering in extreme proximity around TINs and, to a lesser extent, Bystander neurons. In addition, we found that T. gondii transcripts were primarily found in the TIN transcriptome, not in the Bystander transcriptome. Collectively, these data suggest that, contrary to common perception, neurons that directly interact with or harbor parasites can be recognized by CD8+ T cells. IMPORTANCE Like other persistent intracellular pathogens, Toxoplasma gondii, a protozoan parasite, has evolved to evade the immune system and establish a chronic infection in specific cells and organs, including neurons in the CNS. Understanding T. gondii’s persistence in neurons holds the potential to identify novel, curative drug targets. The work presented here offers new insights into the neuron-T. gondii interaction in vivo. By transcriptionally profiling neurons manipulated by T. gondii, we unexpectedly revealed that immune cells, and specifically CD8+ T cells, appear to cluster around these neurons, suggesting that CD8+ T cells specifically recognize parasite-manipulated neurons. Such a possibility supports evidence from other labs that questions the long-standing dogma that neurons are often persistently infected because they are not directly recognized by immune cells such as CD8+ T cells. Collectively, these data suggest we reconsider the broader role of neurons in the context of infection and neuroinflammation.

scholarly journals Identification of Mycobacterial RplJ/L10 and RpsA/S1 Proteins as Novel Targets for CD4+ T Cells

2017 ◽  
Vol 85 (4) ◽  
Author(s):  
Alison J. Johnson ◽  
Steven C. Kennedy ◽  
Cecilia S. Lindestam Arlehamn ◽  
Michael F. Goldberg ◽  
Neeraj K. Saini ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Mhc Class Ii ◽  
Cell Response ◽  
Large Subunit ◽  
T Cell Response ◽  
Critical Feature ◽  
Content Type ◽  
Efficacious Vaccine

ABSTRACT Tuberculosis (TB) due to Mycobacterium tuberculosis remains a major global infectious disease problem, and a more efficacious vaccine is urgently needed for the control and prevention of disease caused by this organism. We previously reported that a genetically modified strain of Mycobacterium smegmatis called IKEPLUS is a promising TB vaccine candidate. Since protective immunity induced by IKEPLUS is dependent on antigen-specific CD4+ T cell memory, we hypothesized that the specificity of the CD4+ T cell response was a critical feature of this protection. Using in vitro assays of interferon gamma production (enzyme-linked immunosorbent spot [ELISPOT] assays) by splenocytes from IKEPLUS-immunized C57BL/6J mice, we identified an immunogenic peptide within the mycobacterial ribosomal large subunit protein RplJ, encoded by the Rv0651 gene. In a complementary approach, we generated major histocompatibility complex (MHC) class II-restricted T cell hybridomas from IKEPLUS-immunized mice. Screening of these T cell hybridomas against IKEPLUS and ribosomes enriched from IKEPLUS suggested that the CD4+ T cell response in IKEPLUS-immunized mice was dominated by the recognition of multiple components of the mycobacterial ribosome. Importantly, CD4+ T cells specific for mycobacterial ribosomes accumulate to significant levels in the lungs of IKEPLUS-immunized mice following aerosol challenge with virulent M. tuberculosis, consistent with a role for these T cells in protective host immunity in TB. The identification of CD4+ T cell responses to defined ribosomal protein epitopes expands the range of antigenic targets for adaptive immune responses to M. tuberculosis and may help to inform the design of more effective vaccines against tuberculosis.

scholarly journals Role of CD4 in thymocyte selection and maturation.

1989 ◽  
Vol 169 (6) ◽  
pp. 2085-2096 ◽  
Author(s):  
J C Zuñiga-Pflücker ◽  
S A McCarthy ◽  
M Weston ◽  
D L Longo ◽  
A Singer ◽  
...  
Keyword(s):  
T Cells ◽  
Mhc Class Ii ◽  
Cd8 T Cells ◽  
Decisive Role ◽  
Class Ii ◽  
Double Positive ◽  
Adult Mice

We examined the possible role of CD4 molecules during in vivo and in vitro fetal thymic development. Our results show that fetal thymi treated with intact anti-CD4 mAbs fail to generate CD4 single-positive T cells, while the generation of the other phenotypes remains unchanged. Most importantly, the use of F(ab')2 and Fab anti-CD4 mAb gave identical results, i.e., failure to generate CD4+/CD8- T cells, with no effect on the generation of CD4+/CD8+ T cells. Since F(ab')2 and Fab anti-CD4 fail to deplete CD4+/CD8- in adult mice, these results strongly argue that the absence of CD4+/CD8- T cells is not due to depletion, but rather, is caused by a lack of positive selection, attributable to an obstructed CD4-MHC class II interaction. Furthermore, we also observed an increase in TCR/CD3 expression after anti-CD4 (divalent or monovalent) mAb treatment. The TCR/CD3 upregulation occurs in the double-positive population, and may result from CD4 signaling after mAb engagement, or may be a consequence of the blocked CD4-class II interactions. One proposed model argues that the CD3 upregulation occurs in an effort to compensate for the reduction in avidity or signaling that is normally provided by the interaction of the CD4 accessory molecule and its ligand. As a whole, our findings advocate that CD4 molecules play a decisive role in the differentiation of thymocytes.

scholarly journals Reassessment of the Role of Aromatic Amino Acid Hydroxylases and the Effect of Infection by Toxoplasma gondii on Host Dopamine

2014 ◽  
Vol 83 (3) ◽  
pp. 1039-1047 ◽  
Author(s):  
Zi T. Wang ◽  
Steve Harmon ◽  
Karen L. O'Malley ◽  
L. David Sibley
Keyword(s):  
Amino Acid ◽  
Toxoplasma Gondii ◽  
Aromatic Amino Acid ◽  
Content Type ◽  
Aromatic Amino Acid Hydroxylase ◽  
Aromatic Amino Acid Hydroxylases ◽  
Host Brain

Toxoplasma gondiiinfection has been described previously to cause infected mice to lose their fear of cat urine. This behavioral manipulation has been proposed to involve alterations of host dopamine pathways due to parasite-encoded aromatic amino acid hydroxylases. Here, we report successful knockout and complementation of the aromatic amino acid hydroxylaseAAH2gene, with no observable phenotype in parasite growth or differentiationin vitroandin vivo. Additionally, expression levels of the two aromatic amino acid hydroxylases were negligible both in tachyzoites and in bradyzoites. Finally, we were unable to confirm previously described effects of parasite infection on host dopamine eitherin vitroorin vivo, even whenAAH2was overexpressed using theBAG1promoter. Together, these data indicate that AAH enzymes in the parasite do not cause global or regional alterations of dopamine in the host brain, although they may affect this pathway locally. Additionally, our findings suggest alternative roles for theAHHenzymes inT. gondii, sinceAAH1is essential for growth in nondopaminergic cells.

scholarly journals Succinylated Wheat Germ Agglutinin Colocalizes with the Toxoplasma gondii Cyst Wall Glycoprotein CST1

mSphere ◽  
2020 ◽  
Vol 5 (2) ◽  
Author(s):  
Rebekah B. Guevara ◽  
Barbara A. Fox ◽  
David J. Bzik
Keyword(s):  
Toxoplasma Gondii ◽  
Cyst Wall ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Parasitophorous Vacuole ◽  
Biological Information ◽  
Toxoplasmic Encephalitis ◽  
Content Type ◽  
Cyst Development

ABSTRACT The glycosylated mucin domain of the Toxoplasma gondii cyst wall glycoprotein CST1 is heavily stained by Dolichos biflorus agglutinin, a lectin that binds to N-acetylgalactosamine. The cyst wall is also heavily stained by the chitin binding lectin succinylated wheat germ agglutinin (s-WGA), which selectively binds to N-acetylglucosamine-decorated structures. Here, we tracked the localization of N-acetylglucosamine-decorated structures that bind to s-WGA in immature and mature in vitro cysts. s-WGA localization was observed at the cyst periphery 6 h after the differentiation of the tachyzoite-stage parasitophorous vacuole. By day 1 and at all later times after differentiation, s-WGA was localized in a continuous staining pattern at the cyst wall. Coinciding with the maturation of the cyst matrix by day 3 of cyst development, s-WGA also localized in a continuous matrix pattern inside the cyst. s-WGA localized in both the outer and inner layer regions of the cyst wall and in a continuous matrix pattern inside mature 7- and 10-day-old cysts. In addition, s-WGA colocalized in the cyst wall with CST1, suggesting that N-acetylglucosamine- and N-acetylgalactosamine-decorated molecules colocalized in the cyst wall. In contrast to CST1, GRA4, and GRA6, the relative accumulation of the molecules that bind s-WGA in the cyst wall was not dependent on the expression of GRA2. Our results suggest that GRA2-dependent and GRA2-independent mechanisms regulate the trafficking and accumulation of glycosylated molecules that colocalize in the cyst wall. IMPORTANCE Chronic Toxoplasma gondii infection is maintained in the central nervous system by thick-walled cysts. If host immunity wanes, cysts recrudesce and cause severe and often lethal toxoplasmic encephalitis. Currently, there are no therapies to eliminate cysts, and little biological information is available regarding cyst structure(s). Here, we investigated cyst wall molecules recognized by succinylated wheat germ agglutinin (s-WGA), a lectin that specifically binds to N-acetylglucosamine-decorated structures. N-Acetylglucosamine regulates cell signaling and plays structural roles at the cell surface in many organisms. The cyst wall and cyst matrix were heavily stained by s-WGA in mature cysts and were differentially stained during cyst development. The relative accumulation of molecules that bind to s-WGA in the cyst wall was not dependent on the expression of GRA2. Our findings suggest that glycosylated cyst wall molecules gain access to the cyst wall via GRA2-dependent and GRA2-independent mechanisms and colocalize in the cyst wall.

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