Cell-autonomous Hedgehog signaling controls Th17 polarization and pathogenicity.

Th17 cells are key drivers of autoimmune disease. However, the signaling pathways regulating Th17 polarization are poorly understood. Hedgehog signaling regulates cell fate decisions during embryogenesis and adult tissue patterning. Here we find that cell-autonomous Hedgehog signaling, independent of exogenous ligands, selectively drives the polarization of Th17 cells but not other T helper cell subsets. We show that endogenous Hedgehog ligand, Ihh, signals to activate both canonical and non-canonical Hedgehog pathways through Gli3 and AMPK. We demonstrate that Hedgehog pathway inhibition with either the clinically-approved small molecule inhibitor vismodegib or genetic ablation of Ihh in CD4+ T cells greatly diminishes disease severity in two mouse models of intestinal inflammation. We confirm that Hedgehog pathway expression is upregulated in tissue from human ulcerative colitis patients and correlates with Th17 marker expression. This work implicates Hedgehog signaling in Th17 polarization and intestinal immunopathology and indicates the potential therapeutic use of Hedgehog inhibitors in the treatment of inflammatory bowel disease.

IL-7-dependent compositional changes within the γδ T cell pool in lymph nodes during ageing lead to an unbalanced anti-tumour response.

How the age-associated decline of immune function leads to increased cancer incidence is poorly understood. Here, we have characterised the cellular composition of the γδ T-cell pool in peripheral lymph nodes (pLNs) upon ageing. We find that ageing has minimal cell-intrinsic effects on function and global gene expression of γδ T cells, and γδTCR diversity remains stable. However, ageing alters TCRδ chain usage and clonal structure of γδ T-cell subsets. Importantly, IL-17-producing γδ17 T cells dominate the γδ T-cell pool of aged mice-mainly due to the selective expansion of Vγ6+ γδ17 T cells and augmented γδ17 polarisation of Vγ4+ T cells. Expansion of the γδ17 T-cell compartment is mediated by increased IL-7 expression in the T-cell zone of old mice. In a Lewis lung cancer model, pro-tumourigenic Vγ6+ γδ17 T cells are exclusively activated in the tumour-draining LN and their infiltration into the tumour correlates with increased tumour size in aged mice. Thus, upon ageing, substantial compositional changes in γδ T-cell pool in the pLN lead to an unbalanced γδ T-cell response in the tumour that is associated with accelerated tumour growth.

PI3Kδ hyper-activation promotes development of B cells that exacerbate Streptococcus pneumoniae infection in an antibody-independent manner.

Streptococcus pneumoniae is a major cause of pneumonia and a leading cause of death world-wide. Antibody-mediated immune responses can confer protection against repeated exposure to S. pneumoniae, yet vaccines offer only partial protection. Patients with Activated PI3Kδ Syndrome (APDS) are highly susceptible to S. pneumoniae. We generated a conditional knock-in mouse model of this disease and identify a CD19+B220- B cell subset that is induced by PI3Kδ signaling, resides in the lungs, and is correlated with increased susceptibility to S. pneumoniae during early phases of infection via an antibody-independent mechanism. We show that an inhaled PI3Kδ inhibitor improves survival rates following S. pneumoniae infection in wild-type mice and in mice with activated PI3Kδ. These results suggest that a subset of B cells in the lung can promote the severity of S. pneumoniae infection, representing a potential therapeutic target.

Ionic immune suppression within the tumour microenvironment limits T cell effector function.

Tumours progress despite being infiltrated by tumour-specific effector T cells. Tumours contain areas of cellular necrosis, which are associated with poor survival in a variety of cancers. Here, we show that necrosis releases intracellular potassium ions into the extracellular fluid of mouse and human tumours, causing profound suppression of T cell effector function. Elevation of the extracellular potassium concentration ([K+]e) impairs T cell receptor (TCR)-driven Akt-mTOR phosphorylation and effector programmes. Potassium-mediated suppression of Akt-mTOR signalling and T cell function is dependent upon the activity of the serine/threonine phosphatase PP2A. Although the suppressive effect mediated by elevated [K+]e is independent of changes in plasma membrane potential (Vm), it requires an increase in intracellular potassium ([K+]i). Accordingly, augmenting potassium efflux in tumour-specific T cells by overexpressing the potassium channel Kv1.3 lowers [K+]i and improves effector functions in vitro and in vivo and enhances tumour clearance and survival in melanoma-bearing mice. These results uncover an ionic checkpoint that blocks T cell function in tumours and identify potential new strategies for cancer immunotherapy.

Cowden's syndrome with immunodeficiency.

BACKGROUND: Cowden's syndrome is a rare, autosomal dominant disease caused by mutations in the phosphoinositide 3-kinase and phosphatase and tensin homolog (PTEN) gene. It is associated with hamartomatous polyposis of the gastrointestinal tract, mucocutaneous lesions and increased risk of developing certain types of cancer. In addition to increased risk of tumour development, mutations in PTEN have also been associated with autoimmunity in both mice and humans. Until now, however, an association between Cowden's syndrome and immune deficiency has been reported in a single patient only. METHODS AND RESULTS: Two patients with Cowden's syndrome and an increased frequency of infections were investigated for possible underlying immunodeficiency. In one patient, hypogammaglobulinaemia with a functional antibody deficiency was identified, while the other patient had a persisting CD4+ T cell lymphopenia (with normal antibody production). CONCLUSIONS: Our data indicate that Cowden's syndrome may be associated with both T cell and B cell immune dysfunction. We recommend that patients with Cowden's syndrome and an increased frequency of infections are investigated for associated immunodeficiency.

PI3Kδ Regulates the Magnitude of CD8+ T Cell Responses after Challenge with Listeria monocytogenes.

PI3Ks regulate diverse immune cell functions by transmitting intracellular signals from Ag, costimulatory receptors, and cytokine receptors to control cell division, differentiation, survival, and migration. In this study, we report the effect of inhibiting the p110δ subunit of PI3Kδ on CD8(+) T cell responses to infection with the intracellular bacteria Listeria monocytogenes. A strong dependency on PI3Kδ for IFN-γ production by CD8(+) T cells in vitro was not recapitulated after Listeria infection in vivo. Inactivation of PI3Kδ resulted in enhanced bacterial elimination by the innate immune system. However, the magnitudes of the primary and secondary CD8 +: T cell responses were reduced. Moreover, PI3Kδ activity was required for CD8(+) T cells to provide help to other responding CD8(+) cells. These findings identify PI3Kδ as a key regulator of CD8(+) T cell responses that integrates extrinsic cues, including those from other responding cells, to determine the collective behavior of CD8(+) T cell populations responding to infection.

Metabolism modifications and apoptosis induction after Cellfood™ administration to leukemia cell lines.

BACKGROUND: Cellfood™ (CF) is a nutritional supplement containing deuterium sulphate, minerals, amino acids, and enzymes, with well documented antioxidant properties. Its organic and inorganic components are extracted from the red algae Lithothamnion calcareum, whose mineral extract has shown growth-inhibitory effect both on in vitro and in vivo models. The purpose of this study was to evaluate the antiproliferative effects of CF on leukemic cells. In fact, according to its capacity to modulate O2 availability and to improve mitochondrial respiratory metabolism, we wondered if CF could affect cancer cell metabolism making cells susceptible to apoptosis. METHODS: Three leukemic cell lines, Jurkat, U937, and K562, were treated with CF 5 µl/ml up to 72 hours. Cell viability, apoptosis (i.e. caspase-3 activity and DNA fragmentation), hypoxia inducible factor 1 alpha (HIF-1α) concentration, glucose transporter 1 (GLUT-1) expression, lactate dehydrogenase (LDH) activity and lactate release in the culture medium were detected and compared with untreated cells. RESULTS: CF significantly inhibited leukemic cell viability by promoting cell apoptosis, as revealed by caspase-3 activation and DNA laddering. In particular, CF treated cells showed lower HIF-1α levels and lower GLUT-1 expression as compared to untreated cells. At the same time, CF was able to reduce LDH activity and, consequently, the amount of lactate released in the extracellular environment. CONCLUSIONS: We supplied evidence for an antiproliferative effect of CF on leukemia cell lines by inducing cell death through an apoptotic mechanism and by altering cancer cell metabolism through HIF-1α and GLUT-1 regulation. Thanks to its antioxidative and proapoptotic properties, CF might be a good candidate for cancer prevention.