Search

Hepatic macrophages maintain liver homeostasis, but little is known about the signals that activate the hepatoprotective programs within macrophages. Here, we show that toll-like receptor 13 (TLR13), a sensor of bacterial 23S ribosomal RNA (rRNA), senses microbiome RNAs to drive tissue-protective responses in CD5Lhihepatic macrophages. Splenomegaly and hepatomegaly developed in the absence of the endosomal RNase, RNaseT2, via TLR13-dependent macrophage proliferation. Furthermore, TLR13 in hepatic Ly6Clomacrophages activated the transcription factors LXRα and MafB, leading to expression of tissue-clearance molecules, such as CD5L, C1qb, and Axl. Consequently,Rnaset2−/−mice developed resistance to acute liver injury caused by challenges with acetaminophen and lipopolysaccharide + D-galactosamine. TLR13 responses inRnaset2−/−mice were impaired by antibiotics, suggesting that TLR13 were activated by microbiome rRNAs, which was detected in the sera and hepatic macrophages. Repeated administration of wild-type mice with the TLR13 ligand, rather than other TLR ligands, selectively increased the number of Kupffer cells, which expressed immunoregulatory and tissue-clearance genes as hepatic macrophages inRnaset2−/−mice did. Our results suggest that microbiome ssRNA serves as an environmental cue for initiating tissue-protective TLR13 responses in hepatic macrophages.Graphical AbstractIn the absence of an endosomal RNase, RNase T2, microbiome RNAs circulating in the vasculature activate TLR13 in hepatic macrophages to drive hepatoprotective responses through expression of immunoregulatory and tissue-clearance molecules. Consequently, mice lacking RNase T2 are resistant against acute liver injuries caused by acetaminophen and LPS + D-galactosamine.

SLC29A3, also known as ENT3, is a lysosomal transmembrane protein that transports nucleosides from the lysosomes to the cytoplasm1. Loss-of-function mutations inSLC29A3cause lysosomal nucleoside storage and histiocytosis: phagocyte accumulation in multiple organs2,3. However, little is known about the mechanism through which lysosomal nucleoside storage drives histiocytosis. Herein, histiocytosis inSlc29a3−/−mice was demonstrated to depend on TLR7, which senses a combination of nucleosides and oligoribonucleotides4,5. TLR7 responded to lysosomal nucleoside storage and enhanced proliferation of Ly6ChiCX3CR1lowimmature monocytes and their maturation into Ly6Clowphagocytes inSlc29a3−/−mice. Because accumulated nucleosides primarily originated from cell corpse phagocytosis, TLR7 in immature monocytes recognized nucleoside storage as lysosomal stress and increased phagocyte numbers. This non-inflammatory compensatory response is referred to as the TLR7 stress response where Syk, GSK3β, β-catenin, and mTORC1 serve as downstream signalling molecules. In SLC29A3 disorders, histiocytosis accompanies inflammation6,7. Nucleoside storage failed to induce pro-inflammatory cytokine production inSlc29a3−/−mice, but enhanced ssRNA-dependent pro-inflammatory cytokine production in Ly6Chiclassical monocytes and peripheral macrophages, not proliferating immature monocytes. Patient-derived monocytes harbouring G208RSLC29A3mutation showed higher survival and proliferation in the presence of M-CSF and produced larger amounts of IL-6 upon ssRNA stimulation than did those derived from healthy subjects. A TLR8 antagonist inhibited the survival/proliferation of patient-derived macrophages. These results demonstrated that TLR7/8 responses to lysosomal nucleoside stress drive SLC29A3 disorders.

Toll-like receptors (TLRs) impact myeloid cell responsiveness to environmental cues such as pathogen components and metabolites. Although TLR protein expression in monocytes and tissue macrophages is thought to be optimized for microenvironments in each tissue, a comprehensive study has not been reported. We here examined protein expression of endogenous TLRs in tissue-resident myeloid cells. Neutrophils in peripheral blood, spleen, liver and lung expressed TLR2, TLR4 and TLR5 in all tissues. Ly6C+ MHC II‒ classical monocytes mature into Ly6C‒ MHC II+ monocyte-derived dendritic cells (moDCs) or Ly6C‒ MHC II‒ patrolling monocytes. These subsets were found in all the tissues studied. TLR2 and TLR4 were displayed on all of these subsets, regardless of location. In contrast, expression of endosomal TLRs did vary with tissues and subsets. moDCs expressed TLR9, but much less TLR7. In contrast, TLR7, not TLR3 or TLR9, was highly expressed in classical and patrolling monocytes. Tissue macrophages such as red pulp macrophages in the spleen, Kupffer cells in the liver, microglia in the brain, alveolar macrophages in the lung and adipose tissue macrophages all expressed TLR2, TLR4 and TLR3. TLR7 was also expressed in these tissue macrophages except Kupffer cells in the liver. TLR9 expression in tissue macrophages was much lower or hard to detect. These results suggest that expression of endosomal TLRs in myeloid cells is influenced by their differentiation status and tissue-specific microenvironments.

Toll-like receptor 8 (TLR8), a sensor for pathogen-derived single-stranded RNA (ssRNA), binds to uridine (Uri) and ssRNA to induce defense responses. We here show that cytidine (Cyd) with ssRNA also activated TLR8 in peripheral blood leukocytes (PBLs) and a myeloid cell line U937, but not in an embryonic kidney cell line 293T. Cyd deaminase (CDA), an enzyme highly expressed in leukocytes, deaminates Cyd to Uri. CDA expression enabled TLR8 response to Cyd and ssRNA in 293T cells. CDA deficiency and a CDA inhibitor both reduced TLR8 responses to Cyd and ssRNA in U937. The CDA inhibitor also reduced PBL response to Cyd and ssRNA. A Cyd analogue, azacytidine, is used for the therapy of myelodysplastic syndrome and acute myeloid leukemia. Azacytidine with ssRNA induced tumor necrosis factor-α expression in U937 and PBLs in a manner dependent on CDA and TLR8. These results suggest that CDA enables TLR8 activation by Cyd or its analogues with ssRNA through deaminating activity. Nucleoside metabolism might impact TLR8 responses in a variety of situations such as the treatment with nucleoside analogues.

The Toll-like receptor 4 (TLR4)/myeloid differentiation factor-2 (MD-2) complex is essential for LPS recognition and induces innate immune responses against Gram-negative bacteria. As activation of TLR4/MD-2 is also critical for the induction of adaptive immune responses, TLR4/MD-2 agonists have been developed as vaccine adjuvants, but their efficacy has not yet been ascertained. Here, we demonstrate that a funiculosin (FNC) variant, FNC-RED, and FNC-RED and FNC derivatives are agonists for both murine and human TLR4/MD-2. FNC-RED induced nuclear factor-κB (NF-κB) activation via murine TLR4/MD-2, whereas FNC had no TLR4/MD-2 stimulatory activity. Biacore analysis revealed that FNC-RED binds to murine TLR4/MD-2 but not murine radioprotective 105 (RP105)/myeloid differentiation factor-1 (MD-1), another LPS sensor. FNC-RED induced CD14-independent expressions of pro-inflammatory cytokines and co-stimulatory molecules in murine macrophages and dendritic cells. In contrast, FNC-RED stimulation was reduced in CD14-dependent LPS responses, including dimerization and internalization of TLR4/MD-2 and IFN-β expression. FNC-RED-induced IL-12p40 production from murine dendritic cells was dependent on NF-κB but not MAPK pathway. In addition, fetal bovine serum augmented lipid A-induced NF-κB activation but blocked FNC-RED-mediated responses. Two synthetic phosphate group-containing FNC-RED and FNC derivatives, FNC-RED-P01 and FNC-P01, respectively, activated human TLR4/MD-2, unlike FNC-RED. Finally, computational analysis revealed that this species-specific activation by FNC-RED and FNC-RED-P01 resulted from differences in electrostatic surface potentials between murine and human TLR4/MD-2. We conclude that FNC-RED and its synthetic derivative represent a novel category of murine and human TLR4/MD-2 agonist.

A lysosomal transmembrane protein SLC29A3 transports nucleosides from lysosomes to the cytoplasm. Loss-of-function mutations of the SLC29A3 gene cause lysosomal nucleoside storage in monocyte/macrophages, leading to their accumulation called histiocytosis in humans and mice. Little is known, however, about a mechanism behind nucleoside-dependent histiocytosis. TLR7, an innate immune sensors for single stranded RNA, bind and respond to nucleosides. We here show that they drive nucleoside-mediated histiocytosis. Patrolling monocyte/macrophages accumulate in the spleen of Slc29a3−/− mice but not Slc29a3−/−Tlr7−/− mice. Accumulated patrolling monocyte/macrophages stored nucleosides derived from cell corpse. TLR7 was recruited to phagosomes and activated as evidenced by TLR7-dependent phagosomal maturation. TLR7 induced hyper-responsiveness to M-CSF in Slc29a3−/− monocyte/macrophages. These results suggest that TLR7 drives histiocytosis in SLC29A3 disorders.One Sentence SummarySLC29A3 disorders are caused by activation of TLR7 with accumulated nucleosides in lysosomes.

Toll-like receptor 8 (TLR8) senses single-stranded RNA (ssRNA) and initiates innate immune responses. TLR8 requires proteolytic cleavage at the loop region (Z-loop) between leucine-rich repeat (LRR) 14 and LRR15 for its activation. However, the molecular basis of Z-loop processing remains unknown. To elucidate the mechanism of Z-loop processing, we performed biochemical and structural studies of how the Z-loop affects the function of TLR8. TLR8 with the uncleaved Z-loop is unable to form a dimer, which is essential for activation, irrespective of the presence of agonistic ligands. Crystallographic analysis revealed that the uncleaved Z-loop located on the ascending lateral face prevents the approach of the dimerization partner by steric hindrance. This autoinhibition mechanism of dimerization by the Z-loop might be occurring in the proteins of the same subfamily, TLR7 and TLR9.

Toll-like Receptor 9 (TLR9) is an innate immune receptor recognizing microbial DNA. TLR9 is also activated by self-derived DNA, such as mitochondrial DNA, in a variety of inflammatory diseases. We show here that TLR9 activation in vivo is controlled by an anti-TLR9 monoclonal Ab (mAb). A newly established mAb, named NaR9, clearly detects endogenous TLR9 expressed in primary immune cells. The mAb inhibited TLR9-dependent cytokine production in vitro by bone marrow-derived macrophages and conventional dendritic cells. Furthermore, NaR9 treatment rescued mice from fulminant hepatitis caused by administering the TLR9 ligand CpGB and D-(+)-galactosamine. The production of proinflammatory cytokines induced by CpGB and D-(+)-galactosamine was significantly impaired by the mAb. These results suggest that a mAb is a promising tool for therapeutic intervention in TLR9-dependent inflammatory diseases.

SUMMARY Objectives The present study was designed to clarify the structure of culture and the three components of attitude in a desirable attitude toward dietary management actions in outpatient haemodialysis patients who are in the maintenance phase of treatment. Methods The participants in the study included nine patients undergoing chronic maintenance haemodialysis who have received guidance related to diet and had good test results. Ethnography, by means of participant observation and semi-structured interviews, was chosen as the research method. Findings Desirable attitude of haemodialysis patients in dietary management actions was found to have a chronological progression in one of the components of attitude: propensity of behaviour. Change in behaviour was influenced by affect and cognition. At the base of the structure of attitude lay three factors: valuing cooking with seasonal ingredients and creating special meals for seasonal occasions; family draws near, shows care and gives support; and belief in information perceived to be good for the health, which was influenced by three components of attitude: affect, cognition, and propensity of behaviour, as well as culture. Conclusion Participants continue to value the food culture that they grew up with, which involves their affect towards, and cognition of, dietary management.

SummaryNatural immunoglobulin M (IgM) is reactive to autoantigens and is believed to be important for autoimmunity. Blood pentameric IgM loaded with antigens forms a large immune complex (IC) that contains various elements, including apoptosis inhibitor of macrophage (AIM). Here we demonstrate that this IgM-AIM association contributes to autoantibody production under obese conditions. In mice fed a high-fat diet, natural IgM increased through B cell TLR4 stimulation. AIM associated with IgM and protected AIM from renal excretion, increasing blood AIM levels along with the obesity-induced IgM augmentation. Meanwhile, the AIM association inhibited IgM binding to the Fcα/μ receptor on splenic follicular dendritic cells, thereby protecting the IgM IC from Fcα/μ receptor-mediated internalization. This supported IgM-dependent autoantigen presentation to B cells, stimulating IgG autoantibody production. Accordingly, in obese AIM-deficient (AIM−/−) mice, the increase of multiple IgG autoantibodies observed in obese wild-type mice was abrogated. Thus, the AIM-IgM association plays a critical role in the obesity-associated autoimmune process.

The small GTPase Arl8b localizes primarily to lysosomes and is involved in lysosomal motility and fusion. Here, we show that Arl8b is required for lysosomal degradation of maternal proteins in the visceral yolk sac endoderm (VYSE), an apical cell layer of the visceral yolk sac, of mouse embryos. The VYSE actively takes up maternal materials from uterine fluid and degrades them in lysosomes to provide breakdown products to the embryo as energy sources. Arl8b gene-trap mice (Arl8b−/−) displayed decreased early embryo body size. The Arl8b−/− VYSE exhibited defective endocytic trafficking to the lysosome and accumulation of maternal proteins such as albumin and immunoglobulin G in late endocytic organelles. Furthermore, Transthyretin-Cre;Arl8bflox/flox mice in which Arl8b was ablated specifically in the VYSE also showed decreased embryo body size, defects in trafficking to the lysosome, and reduction of the free amino acid level in the embryos. Taken together, these results suggest that Arl8b mediates lysosomal degradation of maternal proteins in the VYSE, thereby contributing to mouse embryonic development.

Eosinophilic inflammation is the prominent feature of bronchial asthma, though the importance of eosinophils in the pathogenesis of this disease is controversial. We here established monoclonal antibodies against a newly identified cell surface molecule specifically expressed on mouse eosinophils. Eosinophils were highly purified from small intestine lamina propria and thymus as CD11c(+)Gr1(low)F4/80(+)B220(-) cells. Upon comparative microarray analysis for mRNA expressed in eosinophils and other leukocytes, major facilitator super family domain containing 10 (Mfsd10) was identified as a novel eosinophil-specific cell surface molecule. Hybridomas were established from spleen cells of rats immunized with Mfsd10-introduced Ba/F3 cells. One of three monoclonal antibodies against Mfsd10 displayed selective binding activity against eosinophils recovered in bronchoalveolar lavage fluid of ovalbumin-immunized and -challenged mice. Administration of this antibody in vivo induced a significant reduction of eosinophils recruited in the allergic lungs. Anti-Mfsd10 antibody is useful for investigating the pathophysiological roles of eosinophils with its selective binding and neutralizing activity for mouse eosinophils.

Toll-like receptor (TLR)4/MD-2, a sensor for LPS, delivers the MyD88-dependent signal from the cell surface, then traffics to endolysosomes and delivers the TRIF/TICAM-1-dependent signal. Both signals are thought to be dependent on cell surface TLR4/MD-2. Although TLR4/MD-2 is located also in recycling endosomes, the Golgi apparatus or the endoplasmic reticulum, little is known about a role for intracellular TLR4/MD-2 in LPS responses. We here studied intracellular LPS sensing in macrophages. PRAT4A (protein associated with TLR4 A) is a cochaperone for a general chaperone gp96 and required for cell surface expression of TLR4/MD-2. Cell surface TLR4/MD-2 was undetectable on PRAT4A(-/-) thioglycollate-elicited peritoneal macrophages (P-Macs) and bone marrow-derived macrophages (BM-Macs). LPS responses were all abolished in PRAT4A(-/-) P-Macs, whereas a part of LPS responses remained detectable in PRAT4A(-/-) BM-Macs. Of note, LPS responses in PRAT4A(-/-) BM-Macs were not necessarily dependent on TRIF/TICAM-1 signaling. PRAT4A(-/-) BM-Macs showed unimpaired production of both TRIF/TICAM-1-dependent chemokine RANTES (CCL5) and MyD88-dependent chemokine MCP-1 (CCL2). Moreover, up-regulation of co-stimulatory molecules, CD40 and CD86 was not altered. In contrast, TRIF/TICAM-1-dependent production of type I IFN was profoundly impaired. In response to heat-killed bacteria Escherichia coli, BM-Macs also required PRAT4A-independent TLR4/MD-2 for production of MCP-1 (CCL2) and RANTES (CCL5) and for up-regulation of CD40 and CD86, indicating that intracellular TLR4/MD-2 is able to sense phagocytosed bacteria and activate immune responses. These results demonstrate that intracellular TLR4/MD-2 is responsible for unique set of LPS responses.

Toll-like receptor (TLR) 7and 8 were considered to recognize single-strand RNA (ssRNA) from viruses. Although these receptors also respond to synthetic small chemical ligands, such as CL075 and R848, it remains to be determined whether these receptors sense natural small molecules or not. In the structure of human TLR8 (huTLR8) with ssRNA, there are two ligand-binding sites: one binds a uridine and the other binds an oligoribonucleotide (ORN). This finding demonstrates that huTLR8 recognizes degradation products of ssRNA, suggesting the presence of natural small ligands. We here show that TLR7 works as the sensor for guanosine (G)/2′-deoxyguanosine (dG) in the presence of ORN where ORN strengthens TLR7 interaction with G/dG. In addition, modified nucleosides such as 7-methylguanosine, 8-hydroxyguanosine (8-OHG) and 8-hydroxydeoxyguanosine (8-OHdG) activated TLR7 with ORNs. Importantly, 8-OHdG—a well-known oxidative DNA damage marker with unknown function—induced strong cytokine production comparable to G and dG both in mouse and human immune cells. Although 8-OHdG bound TLR7/ORN with lower affinity than dG did in isothermal titration calorimetry, administered 8-OHdG was metabolically more stable than dG in the serum, indicating that 8-OHdG acts on TLR7 as an endogenous ligand in vivo. To address a role of G analogs in the disease state, we also examined macrophages from Unc93b1D34A/D34A mice, which suffer from TLR7-dependent systemic inflammation, and found that Unc93b1D34A/D34A macrophages showed significantly enhanced response to G alone or 8-OHdG with ORN. In conclusion, our results provide evidence that G, dG, 8-OHG and 8-OHdG are novel endogenous ligands for TLR7.

Low-affinity IgG3 Abs to microbial membranes are important for primary immune defense against microbes, but little is known about the importance of TLRs in their production. IgG3 levels were extremely low in mice lacking radioprotective 105 (RP105), a B cell surface molecule structurally related to TLRs. RP105−/− B cells proliferated poorly in response to not only the TLR4 ligand LPS but also TLR2 ligand lipoproteins, both of which mediate the immunostimulatory activity of microbial membranes. RP105−/− mice were severely impaired in hapten-specific Ab production against LPS or lipoproteins. CD138 (syndecan-1)-positive plasma cells were detected after lipid A injection in wild-type spleen but much less in RP105−/− spleen. RP105 ligation in vivo induced plasma cell differentiation. RP105 expression was ∼3-fold higher on marginal zone B cells than on follicular and B1 cells and was down-regulated on germinal center cells. These results demonstrate that a signal via RP105 is uniquely important for regulating TLR-dependent Ab production to microbial membranes.

TLR3 senses viral dsRNA in endolysosomes. The TLR3 ectodomain is cleaved by proteases such as cathepsins in endolysosomes. It remains controversial whether the N-terminal fragment of TLR3 ectodomain (TLR3N) is cleaved off or remains associated with the C-terminal TLR3 fragment (TLR3C). In addition to endosomes, TLR3 is reported to be expressed on the surface of human fibroblasts, but not of human monocyte-derived dendritic cells. Less is known about roles of TLR3N and cell surface TLR3 in dsRNA sensing. In this study, we show the cleavage site of the TLR3 ectodomain and cell surface expression of TLR3 on mouse primary immune cells. TLR3C, which started at 343S, was associated with TLR3N. Both TLR3N and TLR3C were required for activation of IFN-β and NF-κB promoters by dsRNA, demonstrating that dsRNA is sensed by the TLR3N+C complex. Newly established mAbs to mouse TLR3 revealed that cell surface TLR3 was highly expressed on splenic CD8+ dendritic cells and marginal zone B cells. Cell surface expression of TLR3 on these cells was dependent on the TLR-specific transporter Unc93B1. Although cell surface TLR3 was only weakly expressed on macrophages, TLR3 mAb specifically enhanced TLR3 responses to dsRNA. These results demonstrate that dsRNA is sensed by the TLR3N+C complex and that cell surface TLR3 is a promising target for modulating TLR3 responses.

Toll-like receptors (TLRs) recognize a variety of microbial products and induce defense responses. Pathogen sensing by TLRs occurs either on the cell surface or in endolysosomes. TLR-dependent responses are greatly influenced by the site of pathogen sensing. TLR heterodimers TLR1/TLR2 and TLR2/TLR6 recognize tri- or diacylated microbial lipopeptides, respectively. Although TLR1, 2 and 6 are believed to localize on the cell surface of immune cells, little is known about where lipopeptides are signaled. In this study, we established mAbs to TLR1, 2 and 6. TLR1, 2 and 6 were expressed on the surface of B cells, monocytes and dendritic cells in a manner dependent on a TLR-specific chaperone PRAT4A (protein associated with TLR4 A). Cell surface localization of TLR1 or TLR6 was not necessarily required for TLR2 response. Furthermore, a dynamin inhibitor ‘Dynasore’ abolished the lipopeptide responses by preventing lipopeptide internalization into LAMP-1 and LAMP-2 positive compartments. Our findings suggest that lipopeptides elicit TLR1/2 and TLR2/6 signaling in the endolysosomes, but not on the cell surface.

Toll-like receptor 7 (TLR7) senses microbial-derived RNA but can also potentially respond to self-derived RNA. To prevent autoimmune responses, TLR7 is thought to localize in endolysosomes. Contrary to this view, we show here that TLR7 is present on the cell surface of immune cells and that TLR7 responses can be inhibited by an anti-TLR7 antibody. The anti-TLR7 antibody is internalized with TLR7 and accumulates in endolysosomes as an immune complex. TLR7 responses in dendritic cells, macrophages and B cells are all inhibited by the anti-TLR7 antibody. Furthermore, the anti-TLR7 antibody inhibits in vivo cytokine production induced by a TLR7 ligand. Spontaneous TLR7 activation in Unc93b1(D34A/D34A) mice causes lethal inflammation. Progressive inflammation such as splenomegaly, thrombocytopenia and chronic active hepatitis are ameliorated by anti-TLR7 antibody treatment. These results demonstrate that cell surface TLR7 is a promising target for therapeutic intervention in autoimmune diseases.

The present study was designed to clarify the structure of culture and the three components of attitude in a desirable attitude toward dietary management actions in outpatient haemodialysis patients who are in the maintenance phase of treatment.The participants in the study included nine patients undergoing chronic maintenance haemodialysis who have received guidance related to diet and had good test results. Ethnography, by means of participant observation and semi-structured interviews, was chosen as the research method.Desirable attitude of haemodialysis patients in dietary management actions was found to have a chronological progression in one of the components of attitude: propensity of behaviour. Change in behaviour was influenced by affect and cognition. At the base of the structure of attitude lay three factors: valuing cooking with seasonal ingredients and creating special meals for seasonal occasions; family draws near, shows care and gives support; and belief in information perceived to be good for the health, which was influenced by three components of attitude: affect, cognition, and propensity of behaviour, as well as culture.Participants continue to value the food culture that they grew up with, which involves their affect towards, and cognition of, dietary management.

Toll-like receptor 5 (TLR5), a sensor for bacterial flagellin, mounts innate and adaptive immune responses, and has been implicated in infectious diseases, colitis and metabolic syndromes. Although TLR5 is believed to belong to cell surface TLRs, cell surface expression has never been verified. Moreover, it has remained unclear which types of immune cells express TLR5 and contribute to flagellin-dependent responses. In this study we established an anti-mouse TLR5 monoclonal antibody and studied the cell surface expression of TLR5 on immune cells. The macrophage cell line J774 expressed endogenous TLR5 on the cell surface and produced IL-6 and G-CSF in response to flagellin. Cell surface expression of TLR5 and flagellin-induced responses were completely abolished by silencing a TLR-specific chaperone protein associated with TLR4 A (PRAT4A), demonstrating that TLR5 is another client of PRAT4A. In the in vivo immune cells, cell surface TLR5 was mainly found on neutrophils and CD11b (hi) Ly6C (hi) classical monocytes in the bone marrow, circulation, spleen and inflammatory lesions. Ly6C (hi) classical monocytes, but not neutrophils, produced cytokines in response to flagellin. Splenic CD8 (-) CD4 (+) conventional dendritic cells and CD11c (hi) CD11b (hi) lamina propria DCs, also clearly expressed cell surface TLR5. Collectively, cell surface expression of TLR5 is dependent on PRAT4A and restricted to neutrophils, classical monocytes and specific DC subsets.

SummaryBackground The chemokine receptor CCR4 has been implicated in Th2 cell-mediated immune responses. However, other T cell subsets are also known to participate in allergic inflammation. Objective The role of CCR4 in Th1, Th2, and Th17 cell-mediated allergic airway inflammation was investigated. Method We generated an allergic airway inflammation model by adoptive transfer of in vitro-polarized ovalbumin (OVA)-specific Th1, Th2, and Th17 cells. The effect of a low-molecular weight CCR4 antagonist, Compound 22, on this model was examined. Results Upon in vitro polarization of DO11.10 naive T cells, Th1- and Th2-polarized cells dominantly expressed CXCR3 and CCR4, respectively, while Th17-polarized cells expressed CCR6 and CCR4. Intranasal OVA-challenge of mice transferred with each T cell subset induced accumulation of T cells in the lungs. Eosinophils were also massively accumulated in Th2-transferred mice, whereas neutrophils were preferentially recruited in Th1- and Th17-transferred mice. Compound 22, as well as anti-CCL17 or anti-CCL22 antibody selectively suppressed accumulation of Th2 cells and eosinophils in the lungs of Th2-transferred and OVA-challenged mice. Compound 22 also inhibited bronchial hyperresponsiveness but had little effect on goblet cell hyperplasia in Th2-transferred and OVA-challenged mice. Conclusions and Clinical Relevance There were notable differences in allergic lung inflammation mediated by different T cell subsets. CCR4 blockage was selectively effective for suppression of Th2-mediated allergic inflammation by blocking infiltration of Th2 cells.

Background: To elucidate the usefulness of antigen-specific immunotherapy based on oral vaccination with an edible part of the plant, we examined the effect of transgenic (Tg) rice seeds expressing an immunodominant fragment of the group 1 antigen of Dermatophagoides pteronyssinus (Der p 1) on a murine model of asthma. Methods: Mice were orally vaccinated with the Tg or non-Tg rice seeds for 2 weeks, then they were immunized with recombinant Der p 1 (rDer p 1) and alum intraperitoneally. Antigen-induced immune responses, such as proliferation and cytokine production of CD4+ T cells, antigen-specific serum IgE and IgG, and infiltration of inflammatory cells into the airways were investigated in those mice. Results: The proliferation and Th2 cytokine production of CD4+ T cells in vitro, antigen-specific IgE and IgG synthesis as well as accumulation of eosinophils and lymphocytes into the airways in vivo were significantly inhibited by administration of the Tg rice. Conclusions: These results suggest that the edible vaccines using Tg rice seeds are useful for the treatment of allergic disorders including bronchial asthma.