B cell adaptor for PI3-kinase (BCAP) modulates CD8+ effector and memory T cell differentiation.

CD8+ T cells respond to signals via the T cell receptor (TCR), costimulatory molecules, and immunoregulatory cytokines by developing into diverse populations of effector and memory cells. The relative strength of phosphoinositide 3-kinase (PI3K) signaling early in the T cell response can dramatically influence downstream effector and memory T cell differentiation. We show that initial PI3K signaling during T cell activation results in up-regulation of the signaling scaffold B cell adaptor for PI3K (BCAP), which further potentiates PI3K signaling and promotes the accumulation of CD8+ T cells with a terminally differentiated effector phenotype. Accordingly, BCAP-deficient CD8+ T cells have attenuated clonal expansion and altered effector and memory T cell development following infection with Listeria monocytogenes Thus, induction of BCAP serves as a positive feedback circuit to enhance PI3K signaling in activated CD8+ T cells, thereby acting as a molecular checkpoint regulating effector and memory T cell development.

Bifidobacterium breve MRx0004 protects against airway inflammation in a severe asthma model by suppressing both neutrophil and eosinophil lung infiltration.

Asthma is a phenotypically heterogeneous disease. In severe asthma, airway inflammation can be predominantly eosinophilic, neutrophilic, or mixed. Only a limited number of drug candidates are in development to address this unmet clinical need. Live biotherapeutics derived from the gut microbiota are a promising new therapeutic area. MRx0004 is a commensal Bifidobacterium breve strain isolated from the microbiota of a healthy human. The strain was tested prophylactically and therapeutically by oral gavage in a house dust mite mouse model of severe asthma. A strong reduction of neutrophil and eosinophil infiltration was observed in lung bronchoalveolar lavage fluid following MRx0004 treatment. Peribronchiolar and perivascular immunopathology was also reduced. MRx0004 increased lung CD4+CD44+ cells and CD4+FoxP3+ cells and decreased activated CD11b+ dendritic cells. Cytokine analysis of lung tissue revealed reductions of pro-inflammatory cytokines and chemokines involved in neutrophil migration. In comparison, anti-IL-17 antibody treatment effectively reduced neutrophilic infiltration and increased CD4+FoxP3+ cells, but it induced lung eosinophilia and did not decrease histopathology scores. We have demonstrated that MRx0004, a microbiota-derived bacterial strain, can reduce both neutrophilic and eosinophilic infiltration in a mouse model of severe asthma. This novel therapeutic is a promising next-generation drug for management of severe asthma.

Elevated ACKR2 expression is a common feature of inflammatory arthropathies.

Objectives: Chemokines are essential contributors to leucocyte accumulation at sites of inflammatory pathology. Interfering with chemokine or chemokine receptor function therefore represents a plausible therapeutic option. However, our currently limited understanding of chemokine orchestration of inflammatory responses means that such therapies have not yet been fully developed. We have a particular interest in the family of atypical chemokine receptors that fine-tune, or resolve, chemokine-driven responses. In particular we are interested in atypical chemokine receptor 2 (ACKR2), which is a scavenging receptor for inflammatory CC-chemokines and that therefore helps to resolve in vivo inflammatory responses. The objective of the current study was to examine ACKR2 expression in common arthropathies. Methods: ACKR2 expression was measured by a combination of qPCR and immuno-histochemistry. In addition, circulating cytokine and chemokine levels in patient plasma were assessed using multiplexing approaches. Results: Expression of ACKR2 was elevated on peripheral blood cells as well as on leucocytes and stromal cells in synovial tissue. Expression on peripheral blood leucocytes correlated with, and could be regulated by, circulating cytokines with particularly strong associations being seen with IL-6 and hepatocyte growth factor. In addition, expression within the synovium was coincident with aggregates of lymphocytes, potentially atopic follicles and sites of high inflammatory chemokine expression. Similarly increased levels of ACKR2 have been reported in psoriasis and SSc. Conclusion: Our data clearly show increased ACKR2 in a variety of arthropathies and taking into account our, and others', previous data we now propose that elevated ACKR2 expression is a common feature of inflammatory pathologies.

An analysis of the function and expression of D6 on lymphatic endothelial cells.

The mechanisms by which CC chemokine receptor (CCR)7 ligands are selectively presented on lymphatic endothelium in the presence of inflammatory chemokines are poorly understood. The chemokine-scavenging receptor D6 is expressed on lymphatic endothelial cells (LEC) and contributes to selective presentation of CCR7 ligands by suppressing inflammatory chemokine binding to LEC surfaces. As well as preventing inappropriate inflammatory cell attachment to LECs, D6 is specifically involved in regulating the ability of LEC to discriminate between mature and immature dendritic cells (DCs). D6 overexpression reduces immature DC (iDC) adhesion to LECs, whereas D6 knockdown increases adhesion of iDCs that displace mature DCs. LEC D6 expression is regulated by growth factors, cytokines, and tumor microenvironments. In particular, interleukin-6 and interferon-γ are potent inducers, indicating a preferential role for D6 in inflamed contexts. Expression of the viral interleukin-6 homolog from Kaposi sarcoma-associated herpesvirus is also sufficient to induce significant D6 upregulation both in vitro and in vivo, and Kaposi sarcoma and primary effusion lymphoma cells demonstrate high levels of D6 expression. We therefore propose that D6, which is upregulated in both inflammatory and tumor contexts, is an essential regulator of inflammatory leukocyte interactions with LECs and is required for immature/mature DC discrimination by LECs.

Elevated expression of the chemokine-scavenging receptor D6 is associated with impaired lesion development in psoriasis.

D6 is a scavenging-receptor for inflammatory CC chemokines that are essential for resolution of inflammatory responses in mice. Here, we demonstrate that D6 plays a central role in controlling cutaneous inflammation, and that D6 deficiency is associated with development of a psoriasis-like pathology in response to varied inflammatory stimuli in mice. Examination of D6 expression in human psoriatic skin revealed markedly elevated expression in both the epidermis and lymphatic endothelium in "uninvolved" psoriatic skin (ie, skin that was more than 8 cm distant from psoriatic plaques). Notably, this increased D6 expression is associated with elevated inflammatory chemokine expression, but an absence of plaque development, in uninvolved skin. Along with our previous observations of the ability of epidermally expressed transgenic D6 to impair cutaneous inflammatory responses, our data support a role for elevated D6 levels in suppressing inflammatory chemokine action and lesion development in uninvolved psoriatic skin. D6 expression consistently dropped in perilesional and lesional skin, coincident with development of psoriatic plaques. D6 expression in uninvolved skin also was reduced after trauma, indicative of a role for trauma-mediated reduction in D6 expression in triggering lesion development. Importantly, D6 is also elevated in peripheral blood leukocytes in psoriatic patients, indicating that upregulation may be a general protective response to inflammation. Together our data demonstrate a novel role for D6 as a regulator of the transition from uninvolved to lesional skin in psoriasis.

An investigation of the inflammatory cytokine and chemokine network in systemic sclerosis.

OBJECTIVES: Systemic sclerosis (SSc) is characterised by vasculopathy, an aberrantly activated immune system and excessive extracellular matrix deposition. Inflammatory chemokines control migration of cells to sites of tissue damage; their removal from inflamed sites is essential for resolution of the inflammatory response. The atypical chemokine receptor D6 has a critical role in this physiological balance. To explore potential deregulation of this system in SSc, inflammatory chemokine and D6 expression were compared with that in healthy controls (HC). METHODS: Serum levels of inflammatory mediators were assessed by luminex analysis. Peripheral blood mononuclear cells (PBMCs) were used in molecular and immunocytochemical analysis. Platelet-rich plasma was collected and assessed by western blotting for D6 expression levels. Sex-matched HC were used for comparison. RESULTS: 72 patients with SSc and 30 HC were enrolled in the study. The chemokines MCP-1/CCL2, MIP-1α/CCL3, MIP-1ß/CCL4 and IL-8/CXCL8 were significantly increased in patients with SSc, regardless of disease subtype and phase. Quantitative PCR analysis revealed a significant 10-fold upregulation of D6 transcripts in patients with SSc compared with controls, and this was paralleled by increased D6 protein expression in the PBMCs of patients with SSc. Platelet lysates also showed strong D6 expression in patients with SSc but not in controls. Importantly, high levels of D6 expression correlated with reduced levels of its ligands in serum. CONCLUSIONS: Inflammatory chemokines and the regulatory receptor D6 are significantly upregulated in SSc and high D6 levels are associated with lower systemic chemokine levels, indicating that some patients control systemic chemokine levels using D6. These results suggest that chemokines may represent a therapeutic target in SSc.

Dimeric integrin alpha5beta1 ligands confer morphological and differentiation responses to murine embryonic stem cells.

We present the first report utilizing, and showing the functional relevance of, self-assembling polyvalent ligands specific for integrin alpha5beta1 in murine embryonic stem (mES) cell adhesion. Di, tri and tetrameric 9th-10th type III fibronectin domains (FIII9'10) were used to generate clustered integrin alpha5beta1 ligand surfaces for mES cell culture. Compared to gelatin, FIII9'10 (monomer), FIII9'10-trimer and -tetramer, the FIII9'10-dimer supported the highest number of mES cell colonies. No evidence of domain unfolding upon surface adsorption was found. Colonies appeared disperse with a spread cell morphology unless subdued back to a tight morphology with increasing concentrations of leukemia inhibitory factor (LIF). In the presence of LIF, mES cells adherent to the FIII9'10-dimer showed transient upregulation of Oct-4, the mesodermal transcription factor, Brachyury, and the ectodermal marker, Nestin. However, dual upregulation of Nanog maintained the mES cells in a pluripotent state, confirmed by alkaline phosphatase staining. Therefore, the behavior of mES cells adherent to dimeric integrin alpha5beta1 ligands is a largely morphological phenomenon conferring pro-differentiation signals towards mesodermal and ectodermal lineages. This work will be of interest to cell and tissue engineering groups aiming to control ES cell behavior through integrin ligand presentation and synthetic substrates.