Parasite fate and involvement of infected cells in the induction of CD4+ and CD8+ T cell responses to Toxoplasma gondii

During infection with the intracellular parasite Toxoplasma gondii, the presentation of parasite-derived antigens to CD4+ and CD8+ T cells is essential for long-term resistance to this pathogen. Fundamental questions remain regarding the roles of phagocytosis and active invasion in the events that lead to the processing and presentation of parasite antigens. To understand the most proximal events in this process, an attenuated non-replicating strain of T. gondii (the cpsII strain) was combined with a cytometry-based approach to distinguish active invasion from phagocytic uptake. In vivo studies revealed that T. gondii disproportionately infected dendritic cells and macrophages, and that infected dendritic cells and macrophages displayed an activated phenotype characterized by enhanced levels of CD86 compared to cells that had phagocytosed the parasite, thus suggesting a role for these cells in priming naïve T cells. Indeed, dendritic cells were required for optimal CD4+ and CD8+ T cell responses, and the phagocytosis of heat-killed or invasion-blocked parasites was not sufficient to induce T cell responses. Rather, the selective transfer of cpsII-infected dendritic cells or macrophages (but not those that had phagocytosed the parasite) to naïve mice potently induced CD4+ and CD8+ T cell responses, and conferred protection against challenge with virulent T. gondii. Collectively, these results point toward a critical role for actively infected host cells in initiating T. gondii-specific CD4+ and CD8+ T cell responses

Toxoplasma gondii-Induced Activation of EGFR Prevents Autophagy Protein-Mediated Killing of the Parasite

Toxoplasma gondii resides in an intracellular compartment (parasitophorous vacuole) that excludes transmembrane molecules required for endosome-lysosome recruitment. Thus, the parasite survives by avoiding lysosomal degradation. However, autophagy can re-route the parasitophorous vacuole to the lysosomes and cause parasite killing. This raises the possibility that T. gondii may deploy a strategy to prevent autophagic targeting to maintain the non-fusogenic nature of the vacuole. We report that T. gondii activated EGFR in endothelial cells, retinal pigment epithelial cells and microglia. Blockade of EGFR or its downstream molecule, Akt, caused targeting of the parasite by LC3(+) structures, vacuole-lysosomal fusion, lysosomal degradation and killing of the parasite that were dependent on the autophagy proteins Atg7 and Beclin 1. Disassembly of GPCR or inhibition of metalloproteinases did not prevent EGFR-Akt activation. T. gondii micronemal proteins (MICs) containing EGF domains (EGF-MICs; MIC3 and MIC6) appeared to promote EGFR activation. Parasites defective in EGF-MICs (MIC1 ko, deficient in MIC1 and secretion of MIC6; MIC3 ko, deficient in MIC3; and MIC1-3 ko, deficient in MIC1, MIC3 and secretion of MIC6) caused impaired EGFR-Akt activation and recombinant EGF-MICs (MIC3 and MIC6) caused EGFR-Akt activation. In cells treated with autophagy stimulators (CD154, rapamycin) EGFR signaling inhibited LC3 accumulation around the parasite. Moreover, increased LC3 accumulation and parasite killing were noted in CD154-activated cells infected with MIC1-3 ko parasites. Finally, recombinant MIC3 and MIC6 inhibited parasite killing triggered by CD154 particularly against MIC1-3 ko parasites. Thus, our findings identified EGFR activation as a strategy used by T. gondii to maintain the non-fusogenic nature of the parasitophorous vacuole and suggest that EGF-MICs have a novel role in affecting signaling in host cells to promote parasite survival

Partial-thickness rotator cuff tears: clinical and imaging outcomes and prognostic factors of successful nonoperative treatment

Ian K Lo,1 Matthew R Denkers,2 Kristie D More,1 Atiba A Nelson,1 Gail M Thornton,1 Richard S Boorman1 1Department of Surgery, McCaig Institute for Bone and Joint Health, University of Calgary, Calgary, AB, Canada; 2Division of Orthopaedic Surgery, McMaster University, Hamilton, ON, Canada Purpose: The purpose of this study was to determine the clinical success rate of nonoperative treatment of partial-thickness rotator cuff tears (PT-RCTs), to determine baseline clinical factors predictive of outcome of nonoperative treatment of PT-RCTs, and to determine the imaging outcome of nonoperative treatment of PT-RCTs. Patients and methods: All patients with a primary diagnosis of a PT-RCT were eligible for inclusion. Seventy-six patients (48 males, 28 females) with an average age of 52±10 years were included in the study. Patients were evaluated using a standardized format including clinical, imaging, and shoulder specific quality-of-life outcomes. Patients were assessed and treated either successfully nonoperatively or consented to undergo surgical intervention of their PT-RCT. Patients treated nonoperatively underwent follow-up by MRI arthrogram. Results: Thirty-seven patients (49%) underwent nonoperative treatment. Logistic regression analysis indicated that the baseline variables of side (dominant or nondominant side involved), onset (traumatic or atraumatic), and thickness of tendon tear (<50% or >50%) were significant predictors of outcome. At a mean 46±7 months of follow-up, nonoperatively treated patients demonstrated a mean American Shoulder and Elbow Surgeons score of 85.1±16.0, and a Simple Shoulder Test score of 10.0±2.5. Overall, 76% of tears treated nonoperatively did not show a tear progression on anatomic imaging. Nine patients (24%) demonstrated tear progression, of which three patients (8%) demonstrated full-thickness tearing. Conclusion: Nonoperative treatment was utilized in ~50% of the patients and resulted in improved clinical outcomes. Onset, shoulder involved, and thickness of the tear were predictive of the success of nonoperative treatment. Keywords: magnetic resonance imaging follow-up, nonoperative, partial-thickness rotator cuff tears, rotator cuf

Neurons are the Primary Target Cell for the Brain-Tropic Intracellular Parasite <i>Toxoplasma gondii</i>

<div><p><i>Toxoplasma gondii</i>, a common brain-tropic parasite, is capable of infecting most nucleated cells, including astrocytes and neurons, <i>in vitro</i>. Yet, <i>in vivo</i>, <i>Toxoplasma</i> is primarily found in neurons. <i>In vitro</i> data showing that interferon-γ-stimulated astrocytes, but not neurons, clear intracellular parasites suggest that neurons alone are persistently infected <i>in vivo</i> because they lack the ability to clear intracellular parasites. Here we test this theory by using a novel <i>Toxoplasma</i>-mouse model capable of marking and tracking host cells that directly interact with parasites, even if the interaction is transient. Remarkably, we find that <i>Toxoplasma</i> shows a strong predilection for interacting with neurons throughout CNS infection. This predilection remains in the setting of IFN-γ depletion; infection with parasites resistant to the major mechanism by which murine astrocytes clear parasites; or when directly injecting parasites into the brain. These findings, in combination with prior work, strongly suggest that neurons are not incidentally infected, but rather they are <i>Toxoplasma</i>’s primary <i>in vivo</i> target.</p></div

Generalized Lévy walks and the role of chemokines in migration of effector CD8+ T cells

Chemokines play a central role in regulating processes essential to the immune function of T cells(1-3), such as their migration within lymphoid tissues and targeting of pathogens in sites of inflammation. Here we track T cells using multi-photon microscopy to demonstrate that the chemokine CXCL10 enhances the ability of CD8(+) T cells to control the pathogen T. gondii in the brains of chronically infected mice. This chemokine boosts T cell function in two different ways: it maintains the effector T cell population in the brain and speeds up the average migration speed without changing the nature of the walk statistics. Remarkably, these statistics are not Brownian; rather, CD8(+) T cell motility in the brain is well described by a generalized Lévy walk. According to our model, this surprising feature enables T cells to find rare targets with more than an order of magnitude more efficiency than Brownian random walkers. Thus, CD8(+) T cell behavior is similar to Lévy strategies reported in organisms ranging from mussels to marine predators and monkeys(4-10), and CXCL10 aids T cells in shortening the average time to find rare targets