Chronic infection with؉ T cells does not require their production of interferon-␥, the major mediator to prevent proliferation of tachyzoites during acute infection, but does require perforin. These results suggest that CD8 ؉ T cells induce elimination of T. gondii cysts through their perforin-mediated cytotoxic activity. Our findings provide a new mechanism of the immune system to fight against chronic infection with T. gondii and suggest a possibility of developing a novel vaccine to eliminate cysts from patients with chronic infection and to prevent the establishment of chronic infection after a newly acquired infection.

Notch and Angiopoietin-1 (Ang1)/Tie2 pathways are crucial for vascular maturation and stability. Here we identify the transcription factor ERG as a key regulator of endothelial Notch signalling. We show that ERG controls the balance between Notch ligands by driving Delta-like ligand 4 (Dll4) while repressing Jagged1 (Jag1) expression. In vivo, this regulation occurs selectively in the maturing plexus of the mouse developing retina, where Ang1/Tie2 signalling is active. We find that ERG mediates Ang1-dependent regulation of Notch ligands and is required for the stabilizing effects of Ang1 in vivo. We show that Ang1 induces ERG phosphorylation in a phosphoinositide 3-kinase (PI3K)/Akt-dependent manner, resulting in ERG enrichment at Dll4 promoter and multiple enhancers. Finally, we demonstrate that ERG directly interacts with Notch intracellular domain (NICD) and β-catenin and is required for Ang1-dependent β-catenin recruitment at the Dll4 locus. We propose that ERG coordinates Ang1, β-catenin and Notch signalling to promote vascular stability.

Pericyte investment into new blood vessels is essential for vascular development such that misregulation within this phase of vessel formation can contribute to numerous pathologies including arteriovenous and cerebrovascular malformations. It is critical therefore to illuminate how angiogenic signaling pathways intersect to regulate pericyte migration and investment. Here, we disrupted vascular endothelial growth factor-A (VEGF-A) signaling in ex vivo and in vitro models of sprouting angiogenesis, and found pericyte coverage to be compromised during VEGF-A perturbations. Pericytes had little to no expression of VEGF receptors, suggesting VEGF-A signaling defects affect endothelial cells directly but pericyte indirectly. Live imaging of ex vivo angiogenesis in mouse embryonic skin revealed limited pericyte migration during exposure to exogenous VEGF-A. During VEGF-A gain-of-function conditions, pericytes and endothelial cells displayed abnormal transcriptional changes within the platelet-derived growth factor-B (PDGF-B) and Notch pathways. To further test potential crosstalk between these pathways in pericytes, we stimulated embryonic pericytes with Notch ligands Delta-like 4 (Dll4) and Jagged-1 (Jag1) and found induction of Notch pathway activity but no changes in PDGF Receptor-β (Pdgfrβ) expression. In contrast, PDGFRβ protein levels decreased with mis-regulated VEGF-A activity, observed in the effects on full-length PDGFRβ and a truncated PDGFRβ isoform generated by proteolytic cleavage or potentially by mRNA splicing. Overall, these observations support a model in which, during the initial stages of vascular development, pericyte distribution and coverage are indirectly affected by endothelial cell VEGF-A signaling and the downstream regulation of PDGF-B-PDGFRβ dynamics, without substantial involvement of pericyte Notch signaling during these early stages.

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dReactivation of chronic infection with Toxoplasma gondii can cause life-threatening toxoplasmic encephalitis in immunocompromised individuals. We examined the role of VCAM-1/␣4␤1 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-␣4 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 CD3␦, CD4, CD8␤, gamma interferon (IFN-␥), and inducible nitric oxide synthase (NOS2) (an effector molecule to inhibit T. gondii growth) and the numbers of CD4 ؉ and CD8 ؉ T cells in the brain were significantly less in mice treated with anti-␣4 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-␣4 integrin antibody-treated than in control antibody-treated animals, even though IFN-␥ and NOS2 mRNA levels were higher in the former than in the latter. These results indicate that VCAM-1/␣4␤1 integrin interaction is crucial for prompt recruitment of immune T cells and induction of IFN-␥-mediated protective immune responses during the early stage of reactivation of chronic T. gondii infection to control tachyzoite growth.

Vascular pericytes provide critical contributions to the formation and integrity of the blood vessel wall within the microcirculation. Pericytes maintain vascular stability and homeostasis by promoting endothelial cell junctions and depositing extracellular matrix (ECM) components within the vascular basement membrane, among other vital functions. As their importance in sustaining microvessel health within various tissues and organs continues to emerge, so does their role in a number of pathological conditions including cancer, diabetic retinopathy, and neurological disorders. Here, we review vascular pericyte contributions to the development and remodeling of the microcirculation, with a focus on the local microenvironment during these processes. We discuss observations of their earliest involvement in vascular development and essential cues for their recruitment to the remodeling endothelium. Pericyte involvement in the angiogenic sprouting context is also considered with specific attention to crosstalk with endothelial cells such as through signaling regulation and ECM deposition. We also address specific aspects of the collective cell migration and dynamic interactions between pericytes and endothelial cells during angiogenic sprouting. Lastly, we discuss pericyte contributions to mechanisms underlying the transition from active vessel remodeling to the maturation and quiescence phase of vascular development.

Toxoplasma IgG and IgA, but not IgM, antibody titers were significantly higher in immunocompetent mice with cerebral proliferation of tachyzoites during the chronic stage of infection than those treated with sulfadiazine to inhibit the parasite growth. Their IgG and IgA antibody titers correlated significantly with the amounts of tachyzoite-specific SAG1 mRNA in their brains. In contrast, neither IgG, IgA, nor IgM antibody titers increased following two different doses of challenge infection in chronically infected mice. Increased antibody titers in IgG and IgA but not IgM may be a useful indicator suggesting an occurrence of cerebral tachyzoite growth in immunocompetent individuals chronically infected with T. gondii.