Yoichi Maekawa

Escape of malaria parasites from host immunity requires CD4+ CD25+ regulatory T cells.

Infection with malaria parasites frequently induces total immune suppression, which makes it difficult for the host to maintain long-lasting immunity. Here we show that depletion of CD4+CD25+ regulatory T cells (Treg) protects mice from death when infected with a lethal strain of Plasmodium yoelii, and that this protection is associated with an increased T-cell responsiveness against parasite-derived antigens. These results suggest that activation of Treg cells contributes to immune suppression during malaria infection, and helps malaria parasites to escape from host immune responses.

Delta1-Notch3 Interactions Bias the Functional Differentiation of Activated CD4+ T Cells

Following activation by antigen, naive CD4+ T helper precursor cells execute distinct genetic programs that result in their differentiation toward the type 1 or type 2 helper T cell (Th1 or Th2) phenotype. Although the differentiation and function of these Th subsets has been well studied, little is known about the contribution to these differentiation events of cell surface receptors other than those for soluble cytokines, such as IL-12 or IL-4. Here, we provide direct evidence that the Delta1 interaction with Notch3 on CD4+ T cells transduces signals, promoting development toward the Th1 phenotype. The positive role of Notch signaling in effector cell differentiation was dose dependent, with high levels of stimulation resulting in reduced T cell activation. Our data revealed a clear contribution of Notch pathways to Th1 versus Th2 fate decisions, while also providing insight into another mechanism for inhibition of CD4+ T cell activation.

A mutation in the immunoproteasome subunit PSMB8 causes autoinflammation and lipodystrophy in humans

Proteasomes are multisubunit proteases that play a critical role in maintaining cellular function through the selective degradation of ubiquitinated proteins. When 3 additional β subunits, expression of which is induced by IFN-γ, are substituted for their constitutively expressed counterparts, the structure is converted to an immunoproteasome. However, the underlying roles of immunoproteasomes in human diseases are poorly understood. Using exome analysis, we found a homozygous missense mutation (G197V) in immunoproteasome subunit, β type 8 (PSMB8), which encodes one of the β subunits induced by IFN-γ in patients from 2 consanguineous families. Patients bearing this mutation suffered from autoinflammatory responses that included recurrent fever and nodular erythema together with lipodystrophy. This mutation increased assembly intermediates of immunoproteasomes, resulting in decreased proteasome function and ubiquitin-coupled protein accumulation in the patients tissues. In the patients skin and B cells, IL-6 was highly expressed, and there was reduced expression of PSMB8. Downregulation of PSMB8 inhibited the differentiation of murine and human adipocytes in vitro, and injection of siRNA against Psmb8 in mouse skin reduced adipocyte tissue volume. These findings identify PSMB8 as an essential component and regulator not only of inflammation, but also of adipocyte differentiation, and indicate that immunoproteasomes have pleiotropic functions in maintaining the homeostasis of a variety of cell types.

Genetic variations of Toll-like receptor 9 predispose to systemic lupus erythematosus in Japanese population

Background: Systemic lupus erythematosus (SLE) is characterised by dysregulation of autoreactive lymphocytes and antigen-presenting cells. Signalling through Toll-like receptor 9 (TLR9), a mediator of innate immune responses, has a role in activation of dendritic cells and autoreactive B cells. Objective: To investigate whether TLR9 polymorphisms are associated with an increased risk of SLE. Methods: DNA samples were obtained from 220 Japanese patients with SLE (with >4 American College of Rheumatology criteria for SLE) and 203 controls. The genetic variations of TLR9 were detected by PCR, followed by DNA sequencing. The promoter and enhancer activities of TLR9 were measured by luciferase reporter gene assay. The titres of anti-dsDNA antibodies in sera from control or TLR9-deficient mice were analysed by ELISA. Results: The G allele at position +1174 (located in intron 1 of TLR9) is closely associated with an increased risk of SLE (p = 0.029). Furthermore, patients with SLE tend to have C allele at position −1486 (p = 0.11). Both alleles down regulate TLR9 expression by reporter gene assay. TLR9-deficient mice under a C57BL/6 background possess higher titres of anti-dsDNA serum antibodies than control C57BL/6 mice. Conclusions: These results indicate that the presence of the G allele at position +1174 of TLR9 predisposes humans to an increased risk of SLE. It is speculated that TLR9 normally prevents the development of human SLE.

Notch2 integrates signaling by the transcription factors RBP-J and CREB1 to promote T cell cytotoxicity

The acquisition of cytotoxic effector function by CD8+ T cells is crucial for the control of intracellular infection and tumor invasion. However, it remains unclear which signaling pathways are required for the differentiation of CD8+ cytotoxic T lymphocytes. We show here that Notch2-deficient T cells had impaired differentiation into cytotoxic T lymphocytes. In addition, dendritic cells with lower expression of the Notch ligand Delta-like 1 induced the differentiation of cytotoxic T lymphocytes less efficiently. We found that the intracellular domain of Notch2 interacted with a phosphorylated form of the transcription factor CREB1, and together these proteins bound the transcriptional coactivator p300 to form a complex on the promoter of the gene encoding granzyme B. Our results suggest that the highly regulated, dynamic control of T cell cytotoxicity depends on the integration of Notch2 and CREB1 signals.

Notch signaling drives IL-22 secretion in CD4+ T cells by stimulating the aryl hydrocarbon receptor

CD4+ helper T (Th) cells differentiate toward distinct effector cell lineages characterized by their distinct cytokine expression patterns and functions. Multiple Th cell populations secrete IL-22 that contributes to both protective and pathological inflammatory responses. Although the differentiation of IL-22-producing Th cells is controlled by the aryl hydrocarbon receptor (AhR), little is known about the regulatory mechanisms inducing physiological stimulators for AhR. Here, we show that Notch signaling enhances IL-22 production by CD4+ T cells by a mechanism involving AhR stimulation. Notch-mediated stimulation of CD4+ T cells increased the production of IL-22 even in the absence of STAT3. CD4+ T cells from RBP-J-deficient mice had little ability to produce IL-22 through T cell receptor-mediated stimulation. RBP-J-deficient mice were highly susceptible to the detrimental immunopathology associated with ConA-induced hepatitis with little IL-22 production by CD4+ T cells. Exogenous IL-22 protected RBP-J-deficient mice from ConA-induced hepatitis. Notch signaling promoted production of endogenous stimulators for AhR, which further augmented IL-22 secretion. Our studies identify a Notch–AhR axis that regulates IL-22 expression and fine-tunes immune system control of inflammatory responses.

Nippocystatin, a cysteine protease inhibitor from Nippostrongylus brasiliensis, inhibits antigen processing and modulates antigen-specific immune response.

ABSTRACT During infection, parasites evade the host immune system by modulating or exploiting the immune system; e.g., they suppress expression of major histocompatibility complex class II molecules or secrete cytokine-like molecules. However, it is not clear whether helminths disturb the immune responses of their hosts by controlling the antigen-processing pathways of the hosts. In this study, we identified a new cysteine protease inhibitor, nippocystatin, derived from excretory-secretory (ES) products of an intestinal nematode,Nippostrongylus brasiliensis. Nippocystatin, which belongs to cystatin family 2, consists of 144 amino acids and is secreted as a 14-kDa mature form. In vivo treatment of ovalbumin (OVA)-immunized mice with recombinant nippocystatin (rNbCys) profoundly suppressed OVA-specific proliferation of splenocytes but not non-antigen-specific proliferation of splenocytes. OVA-specific cytokine production was also greatly suppressed in rNbCys-treated mice. Although the serum levels of both OVA-specific immunoglobulin G1 (IgG1) and IgG2a were not affected by rNbCys treatment, OVA-specific IgE was preferentially downregulated in rNbCys-treated mice. In vitro rNbCys inhibited processing of OVA by lysosomal cysteine proteases from the spleens of mice. Mice with anti-nippocystatin antibodies became partially resistant to infection with N. brasiliensis. Based on these findings, N. brasiliensis appears to skillfully evade host immune systems by secreting nippocystatin, which modulates antigen processing in antigen-presenting cells of hosts.

Clinical roles of increased populations of Foxp3+CD4+ T cells in peripheral blood from advanced pancreatic cancer patients.

Objectives: Further metastasis should be avoided in pancreatic cancer (PC) patients for effective surgical treatment. Regulatory T cells (Foxp3+CD4+ T cells including CD4+CD25+ T cells and CD4+CD25− T cells) play important roles in tumor immunity. This study aimed to investigate whether regulatory T cells participate in metastasis. Methods: Peripheral blood was withdrawn from PC patients, as well as healthy volunteer donors as controls. The peripheral blood mononuclear cells (PBMCs) were subjected to FACScan analysis after labeling with anti-CD4, anti-CD25, and anti-Foxp3 antibodies. Tumor markers, including DUPAN2 and CA19-9, surface markers, such as the CD4/CD8 ratio, and the CD57+ cell population were assessed. Clinical stages were classified according to the TNM classification. Results: The Foxp3+CD4+ T-cell population among the PBMCs was significantly increased in PC patients (8.10% ± 4.65%) compared with healthy donors (2.47 ± 0.78%) (P < 0.001). No significant relationships existed for the tumor markers, CD4/CD8 ratio, and CD57+ cells. However, a significant correlation was found between Foxp3+CD4+ T cells among the PBMCs and the TNM stage (P < 0.05). Conclusions: Foxp3+CD4+ T cells are good markers for metastasis detection in PC patients and more accurate than other conventional tumor markers, especially at advanced stages of the disease.

Notch controls the survival of memory CD4+ T cells by regulating glucose uptake

CD4+ T cells differentiate into memory T cells that protect the host from subsequent infection. In contrast, autoreactive memory CD4+ T cells harm the body by persisting in the tissues. The underlying pathways controlling the maintenance of memory CD4+ T cells remain undefined. We show here that memory CD4+ T cell survival is impaired in the absence of the Notch signaling protein known as recombination signal binding protein for immunoglobulin κ J region (Rbpj). Treatment of mice with a Notch inhibitor reduced memory CD4+ T cell numbers and prevented the recurrent induction of experimental autoimmune encephalomyelitis. Rbpj-deficient CD4+ memory T cells exhibit reduced glucose uptake due to impaired AKT phosphorylation, resulting in low Glut1 expression. Treating mice with pyruvic acid, which bypasses glucose uptake and supplies the metabolite downstream of glucose uptake, inhibited the decrease of autoimmune memory CD4+ T cells in the absence of Notch signaling, suggesting memory CD4+ T cell survival relies on glucose metabolism. Together, these data define a central role for Notch signaling in maintaining memory CD4+ T cells through the regulation of glucose uptake.

The lifestyle of the segmented filamentous bacterium: A non-culturable gut-associated immunostimulating microbe inferred by whole-genome sequencing

Numerous microbes inhabit the mammalian intestinal track and strongly impact host physiology; however, our understanding of this ecosystem remains limited owing to the high complexity of the microbial community and the presence of numerous non-culturable microbes. Segmented filamentous bacteria (SFBs), which are clostridia-related Gram-positive bacteria, are among such non-culturable populations and are well known for their unique morphology and tight attachment to intestinal epithelial cells. Recent studies have revealed that SFBs play crucial roles in the post-natal maturation of gut immune function, especially the induction of Th17 lymphocytes. Here, we report the complete genome sequence of mouse SFBs. The genome, which comprises a single circular chromosome of 1 620 005 bp, lacks genes for the biosynthesis of almost all amino acids, vitamins/cofactors and nucleotides, but contains a full set of genes for sporulation/germination and, unexpectedly, for chemotaxis/flagella-based motility. These findings suggest a triphasic lifestyle of the SFB, which comprises two types of vegetative (swimming and epicellular parasitic) phases and a dormant (spore) phase. Furthermore, SFBs encode four types of flagellin, three of which are recognized by Toll-like receptor 5 and could elicit the innate immune response. Our results reveal the non-culturability, lifestyle and immunostimulation mechanisms of SFBs and provide a genetic basis for the future development of the SFB cultivation and gene-manipulation techniques.