Biomedical subjects

Shizuo Akira

Publications and source records attributed to Shizuo Akira.

At least 91 records · Page 5Linked to original sources

PubMed2006

Early type I interferon-mediated signals on B cells specifically enhance antiviral humoral responses.

Type I interferons (IFN-I) limit viral spread by inducing antiviral genes in infected target cells and by shaping the adaptive response through induction of additional cytokines. Vesicular stomatitis virus (VSV) efficiently triggers the production of IFN-I in mice, and it is suggested that IFN-alpha is induced after binding of VSV to TLR7 in infected cells. Our study with virus-specific B cell receptor-transgenic mice demonstrates here that IFN-I directly fuel early humoral immune responses in vivo. VSV-specific B cells that lacked IFN-alpha/beta receptors were considerably impaired in plasma cell formation and in generating antiviral IgM. At low viral titers, production of IFN-alpha following VSV infection was independent of TLR7-mediated signals. Interestingly, however, TLR7 ligation in B cells increased the formation of early antiviral IgM. These findings indicate that IFN-alpha-mediated augmentation of specific B cell responses is a partially TLR7- and virus dose-dependent mechanism.

Animals↗

PubMed2006

TLR2 engagement on CD8 T cells lowers the threshold for optimal antigen-induced T cell activation.

TLR have a crucial role in the detection of microbial infection in mammals. Until recently, most investigations on TLR have focused on cells of the innate immune system and on the role of TLR in the initiation of antigen-specific responses following recognition of microbial products by APC. Here, we report that murine T cells express TLR1, TLR2, TLR6, TLR7 and TLR9 mRNA. Using CD8 T cells from F5 TCR-transgenic mice, we demonstrate that the lipopeptide Pam(3)CysSK(4) (Pam), a synthetic analog of bacterial and mycoplasmal lipoproteins that recognizes TLR1/2 complex, costimulates antigen-activated T cells. Costimulation with Pam permits an increased cell proliferation and survival associated with a sustained CD25 expression and an enhanced expression of Bcl-xL anti-apoptotic protein. In addition, we show that costimulation with Pam up-regulates IFN-gamma production but also granzyme B secretion and cytotoxic activity of antigen-activated T cells, indicating that TLR2 engagement enhances the major effector functions of CD8 T cells. Finally, we demonstrate that TLR2 engagement on T cells lowers the activation threshold for costimulatory signals delivered by APC.

Animals↗

PubMed2006

Nucleic acid agonists for Toll-like receptor 7 are defined by the presence of uridine ribonucleotides.

Toll-like receptor 7 (TLR7) mediates innate responses by responding to viral RNA in endocytic compartments. However, the molecular pattern recognised by TLR7 and whether it differs between RNA of viral and self origin remains unclear. Here, we identify nucleic acids that act as TLR7 agonists for mouse and human cells. We show that uridine and ribose, the two defining features of RNA, are both necessary and sufficient for TLR7 stimulation, and that short single-stranded RNA (ssRNA) act as TLR7 agonists in a sequence-independent manner as long as they contain several uridines in close proximity. Consistent with the notion that TLR7 lacks specificity for sequence motifs, we show that it is triggered equally efficiently by viral or self RNA delivered to endosomes. Our results support the notion that TLR7 recognises uracil repeats in RNA and that it discriminates between viral and self ligands on the basis of endosomal accessibility rather than sequence.

Amino Acid Motifs↗

PubMed2006

Activation of smooth muscle and myenteric plexus cells of jejunum via Toll-like receptor 4.

The cell types of the gut expressing Toll-like receptor 4, which recognizes specifically bacterial lipopolysaccharides, as well as the functionality of this receptor, have remained controversial. We aimed to clarify these issues. Mouse and human intestinal specimens were stained immunohistochemically to detect Toll-like receptor 4 expression. Smooth muscle and myenteric plexus cells but not enterocytes revealed receptor expression. Murine intestinal smooth muscle and myenteric plexus cells but not enterocytes showed nuclear translocation of nuclear factor-kappaB after in vivo stimulation with lipopolysaccharide. Moreover, lipopolysaccharide added to human jejunum biopsies free of epithelial cells induced release of interleukin-8 (IL-8). We can conclude that Toll-like receptor 4 is not expressed in epithelial layer, but rather on smooth muscle and myenteric plexus cells and that expression is functional. The expression of Toll-like receptor 4 on smooth muscle and myenteric plexus cells is consistent with the possibility that these cells are involved in intestinal immune defense; the low or absent expression of Toll-like receptor 4 on enterocytes might explain the intestinal epithelium hyporesponsiveness to the abundance of LPS in the intestinal lumen.

Animals↗

PubMed2006

Toll-like receptors and innate immunity.

The innate immune system is an evolutionally conserved host defense mechanism against pathogens. Innate immune responses are initiated by pattern recognition receptors (PRRs), which recognize specific structures of microorganisms. Among them, Toll-like receptors (TLRs) are capable of sensing organisms ranging from bacteria to fungi, protozoa, and viruses, and play a major role in innate immunity. However, TLRs recognize pathogens either on the cell surface or in the lysosome/endosome compartment. Recently, cytoplasmic PRRs have been identified to detect pathogens that have invaded cytosols. In this review, we focus on the functions of PRRs in innate immunity and their downstream signaling cascades.

Animals↗

PubMed2006

Role of hepatic STAT3 in brain-insulin action on hepatic glucose production.

STAT3 regulates glucose homeostasis by suppressing the expression of gluconeogenic genes in the liver. The mechanism by which hepatic STAT3 is regulated by nutritional or hormonal status has remained unknown, however. Here, we show that an increase in the plasma insulin concentration, achieved either by glucose administration or by intravenous insulin infusion, stimulates tyrosine phosphorylation of STAT3 in the liver. This effect of insulin was mediated by the hormone's effects in the brain, and the increase in hepatic IL-6 induced by the brain-insulin action is essential for the activation of STAT3. The inhibition of hepatic glucose production and of expression of gluconeogenic genes induced by intracerebral ventricular insulin infusion was impaired in mice with liver-specific STAT3 deficiency or in mice with IL-6 deficiency. These results thus indicate that IL-6-STAT3 signaling in the liver contributes to insulin action in the brain, leading to the suppression of hepatic glucose production.

Animals↗

PubMed2006

Toll-like receptors on hematopoietic progenitor cells stimulate innate immune system replenishment.

Toll-like receptors (TLRs) are best known for their ability to recognize microbial or viral components and initiate innate immune responses. We showed here that TLRs and their coreceptors were expressed by multipotential hematopoietic stem cells, whose cell cycle entry was triggered by TLR ligation. TLR expression also extended to some of the early hematopoietic progenitors, although not the progenitor cells dedicated to megakaryocyte and erythroid differentiation. TLR signaling via the Myd88 adaptor protein drove differentiation of myeloid progenitors, bypassing some normal growth and differentiation requirements, and also drove lymphoid progenitors to become dendritic cells. CD14 contributed to the efficiency of lipopolysaccharide (LPS) recognition by stem and progenitor cells, and LPS interacted directly with the TLR4/MD-2 complex on these cells in bone marrow. Thus, the preferential pathogen-mediated stimulation of myeloid differentiation pathways may provide a means for rapid replenishment of the innate immune system during infection.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Toll-like receptor 7-dependent loss of B cell tolerance in pathogenic autoantibody knockin mice.

Systemic lupus erythematosus (SLE) is characterized by the production of autoantibodies that are frequently directed against nucleic acid-associated antigens. To better understand how B cells reactive with such antigens are regulated, we generated a model system in which heavy and light chain genes encoding 564 immunoglobulin have been targeted to the heavy and light chain loci of the nonautoimmune C57BL/6 mouse strain. This antibody recognizes RNA, single-stranded DNA, and nucleosomes. We show that B cells expressing this immunoglobulin were activated, producing class-switched autoantibody in vivo despite the apparently normal induction of anergy. This autoantibody production was largely dependent on Toll-like receptor 7 (TLR7). We further show that production of these autoantibodies was sufficient to cause kidney pathology in these mice. These results demonstrate that the particular threat of nucleic acid-containing autoantigens lies in their ability to bind both antigen receptor and TLR7.

Animals↗

PubMed2006

Innate immune recognition of viral infection.

Induction of the antiviral innate immune response depends on recognition of viral components by host pattern-recognition receptors. Members of the Toll-like receptor family have emerged as key sensors that recognize viral components such as nucleic acids. Toll-like receptor signaling results in the production of type I interferon and inflammatory cytokines and leads to dendritic cell maturation and establishment of antiviral immunity. Cells also express cytoplasmic RNA helicases that function as alternative pattern-recognition receptors through recognition of double-stranded RNA produced during virus replication. These two classes of pattern-recognition receptor molecules are expressed in different intracellular compartments and induce type I interferon responses via distinct signaling pathways.

Adaptor Proteins, Signal Transducing↗

PubMed2006

IL-18 is redundant in T-cell responses and in joint inflammation in antigen-induced arthritis.

IL-18 is an important cofactor in Th1 immune responses and it has additional roles in inflammation. Recent reports suggest the contribution of IL-18 to immune responses may vary between mouse strains and immune contexts. We investigated the contribution of IL-18 to T-cell activation and joint inflammation in Ag-induced arthritis (AIA) in C57Bl/6 mice. AIA and cutaneous delayed-type hypersensitivity (DTH) reactions were induced in wild-type (WT) and IL-18-/- C57Bl/6 mice, and Ag-specific T-cell proliferation and IFN-gamma and IL-4 production were measured. The humoral immune response was measured as serum antibody to the disease-initiating Ag, methylated BSA (mBSA). Splenocyte production of IL-6 was measured by ELISA. To confirm the dependence of this model on Th1-cell-mediated immunity, IL-12p40-/- mice were similarly studied. WT mice developed synovitis, joint effusion, cartilage destruction and bone damage associated with induction of DTH, and in vitro Ag-specific T-cell proliferation and IFN-gamma production. Unexpectedly, IL-18-/- mice developed AIA and indices of T-cell activation were similar to those of WT mice. In contrast, IL-12p40-/- mice did not develop AIA, DTH or T-cell activation. WT and IL-18-/- mice, but not IL-12p40-/- mice, developed significantly increased serum antibody to mBSA compared with naive controls. WT and IL-18-/- splenocytes produced high levels of IL-6, whereas IL-12p40-/- cells had significantly lower IL-6 production compared with both. In conclusion, IL-18 is redundant both as a Th1 response cofactor and inflammatory cytokine, whereas IL-12p40-/- is a key cytokine, in AIA in C57Bl/6 mice.

Animals↗

PubMed2006

Differential inductions of TNF-alpha and IGTP, IIGP by structurally diverse classic and non-classic lipopolysaccharides.

The innate immune system recognizes microbes by characteristic molecules like the Gram-negative lipopolysaccharide (LPS). Lipid A (the LPS bioactive moiety) signals through toll-like receptors (TLRs) to induce pro-inflammatory molecules and small GTPases of the p47 family involved in intracellular pathogen control. We tested TNF-alpha and p47-GTPase induction in macrophages using classical LPSs [lipid As with glucosamine backbones, ester- and amide-linked C14:0(3-OH) and C12 to C16 in acyloxyacyl groups] of wild type and mutant Escherichia coli and Yersinia species and non-classical LPSs [lipid As with diaminoglucose, ester-linked 3-OH-fatty acids and C28:0(27-OH) and C23:0(29-OH) in acyloxyacyl groups] of plant endosymbionts (Rhizobium), intracellular pathogens (Brucella and Legionella) and phylogenetically related opportunistic bacteria (Ochrobactrum). Classical but not non-classical LPSs efficiently induced TNF-alpha, IIGP and IGTP p47-GTPase expression. Remarkably, the acyloxyacyl groups in classical LPSs necessary to efficiently induce TNF-alpha were not necessary to induce p47-GTPases, suggesting that different aspects of lipid A are involved in this differential induction. This was confirmed by using PPDM2, a non-endotoxic lipid A-structurally related synthetic glycolipid. Despite their different bioactivity, all types of LPSs signalled through TLR-4 and not through TLR-2. However, whereas TNF-alpha induction was myeloid differentiation factor 88 (MyD88)-dependent, that of p47-GTPases occurred via a MyD88-independent pathway. These observations show that different aspects of the LPS pathogen-associated molecular pattern may be triggering different signalling pathways linked to the same TLR. They also reinforce the hypothesis that non-classical lipid As act as virulence factors by favouring the escape from the innate immune system.

Animals↗

PubMed2006

The synthetic analogue of mycoplasmal lipoprotein FSL-1 induces dendritic cell maturation through Toll-like receptor 2.

Granulocyte-macrophage colony-stimulating factor-differentiated bone marrow-derived dendritic cells were stimulated with the synthetic lipopeptide S-(2,3-bispalmitoyloxypropyl)-CGDPKHSPKSF (FSL-1) or the Escherichia coli lipopolysaccharide. FSL-1 induced the production of TNF-alpha and IL-12 by C57BL/6-derived bone marrow-derived dendritic cells but not by bone marrow-derived dendritic cells from Toll-like receptor 2-deficient (TLR2(-/-)) mice. Lipopolysaccharide induced the production of TNF-alpha and IL-12 by bone marrow-derived dendritic cells derived from either type of mice. FSL-1 did not induce production of IL-10 by bone marrow-derived dendritic cells from either type of mice, whereas lipopolysaccharide induced small amounts of IL-10 by bone marrow-derived dendritic cells from both types of mice. The upregulation by FSL-1 of the expression of CD80, CD86 and the MHC class II molecule IA(b) was dose- and time-dependent on the surfaces of C57BL/6-derived bone marrow-derived dendritic cells but not on the surface of TLR2(-/-)-derived bone marrow-derived dendritic cells. Lipopolysaccharide upregulated the expression of these molecules on the surfaces of bone marrow-derived dendritic cells from both types of mice. The expression of CD11c on the surfaces of C57BL/6-derived bone marrow-derived dendritic cells was upregulated by stimulation with both FSL-1 and lipopolysaccharide up to 12 h; thereafter, the expression was downregulated. The results suggest that FSL-1 can accelerate maturation of bone marrow-derived dendritic cells and this FSL-1 activity is mediated by TLR2.

Animals↗

PubMed2006

Limited contribution of Toll-like receptor 2 and 4 to the host response to a fungal infectious pathogen, Cryptococcus neoformans.

The present study was designed to elucidate the role of Toll-like receptor (TLR) 2 and TLR4 in the host response to Cryptococcus neoformans. Both TLR2 knockout (KO) and TLR4KO mice produced interleukin-1beta (IL-1beta), IL-6, IL-12p40 and tumor necrosis factor-alpha (TNF-alpha) in sera and cleared this fungal pathogen from infected lungs at a comparable level to control littermate (LM) mice. Synthesis of these cytokines was not significantly different in the lungs of these KO mice and LM mice, although IL-1beta, IL-6 and IL-12p40 tended to be lower in TLR2KO, but not TLR4KO, mice than in controls. In addition, there was no significant reduction detected in the synthesis of IL-12 and TNF-alpha by bone marrow-derived dendritic cells from TLR2KO and TLR4KO mice upon stimulation with live yeast cells. Finally, HEK293 cells expressing either TLR2/dectin-1 or TLR4/MD2/CD14 did not respond to C. neoformans in the activation of nuclear factor kappa B (NFkappaB) detected by a luciferase assay. Our results suggest that TLR2 and TLR4 do not or only marginally contribute to the host and cellular response to this pathogen.

Animals↗

PubMed2006

MyD88 deficiency results in tissue-specific changes in cytokine induction and inflammation in interleukin-18-independent mice infected with Borrelia burgdorferi.

Toll-like receptors (TLRs) play an important role in the control of infection with Borrelia burgdorferi. Deficiencies in TLR-2 or the shared TLR adapter molecule MyD88 have been shown to result in greatly increased bacterial burdens in mice. However, although in vitro studies have shown that the activation of TLR pathways by B. burgdorferi results in the release of inflammatory cytokines, studies in deficient mice have shown either no change or increased rather than decreased inflammation in infected animals. In this study, we looked at mechanisms to explain the increase in inflammation in the absence of MyD88. We found that MyD88-deficient mice infected with B. burgdorferi did not show increased inflammation at sites typically associated with Lyme disease (joints and heart). However, there was markedly increased inflammation in the muscles, kidneys, pancreas, and lungs of deficient animals. Muscle inflammation was typically seen perivascularly and perineuronally similar to that seen in infected humans. Chemotactic chemokines and cytokines were greatly increased in the muscle and kidneys of MyD88-deficient animals but not in the joints or heart tissue, suggesting that MyD88-independent pathways for recognizing B. burgdorferi and inducing these chemokines are present in the muscle and kidneys. Interleukin-18 signaling through MyD88 does not appear to play a role in either control of infection or inflammation.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Toll-like receptor adaptor molecules enhance DNA-raised adaptive immune responses against influenza and tumors through activation of innate immunity.

Toll-like receptors (TLRs) recognize microbial components and trigger the signaling cascade that activates the innate and adaptive immunity. TLR adaptor molecules play a central role in this cascade; thus, we hypothesized that overexpression of TLR adaptor molecules could mimic infection without any microbial components. Dual-promoter plasmids that carry an antigen and a TLR adaptor molecule such as the Toll-interleukin-1 receptor domain-containing adaptor-inducing beta interferon (TRIF) or myeloid differentiation factor 88 (MyD88) were constructed and administered to mice to determine if these molecules can act as an adjuvant. A DNA vaccine incorporated with the MyD88 genetic adjuvant enhanced antigen-specific humoral immune responses, whereas that with the TRIF genetic adjuvant enhanced cellular immune responses. Incorporating the TRIF genetic adjuvant in a DNA vaccine targeting the influenza HA antigen or the tumor-associated antigen E7 conferred superior protection. These results indicate that TLR adaptor molecules can bridge innate and adaptive immunity and potentiate the effects of DNA vaccines against virus infection and tumors.

Adaptor Proteins, Signal Transducing↗

PubMed2006

CCAAT/enhancer-binding protein homologous protein (CHOP) regulates osteoblast differentiation.

Differentiation of committed osteoblasts is controlled by complex activities involving signal transduction and gene expression, and Runx2 and Osterix function as master regulators for this process. Recently, CCAAT/enhancer-binding proteins (C/EBPs) have been reported to regulate osteogenesis in addition to adipogenesis. However, the roles of C/EBP transcription factors in the control of osteoblast differentiation have yet to be fully elucidated. Here we show that C/EBP homologous protein (CHOP; also known as C/EBPzeta) is expressed in bone as well as in mesenchymal progenitors and primary osteoblasts. Overexpression of CHOP reduces alkaline phosphatase activity in primary osteoblasts and suppresses the formation of calcified bone nodules. CHOP-deficient osteoblasts differentiate more strongly than their wild-type counterparts, suggesting that endogenous CHOP plays an important role in the inhibition of osteoblast differentiation. Furthermore, endogenous CHOP induces differentiation of calvarial osteoblasts upon bone morphogenetic protein (BMP) treatment. CHOP forms heterodimers with C/EBPbeta and inhibits the DNA-binding activity as well as Runx2-binding activity of C/EBPbeta, leading to inhibition of osteocalcin gene transcription. These findings indicate that CHOP acts as a dominant-negative inhibitor of C/EBPbeta and prevents osteoblast differentiation but promotes BMP signaling in a cell-type-dependent manner. Thus, endogenous CHOP may have dual roles in regulating osteoblast differentiation and bone formation.

3T3-L1 Cells↗

PubMed2006

BS69, a specific adaptor in the latent membrane protein 1-mediated c-Jun N-terminal kinase pathway.

We previously demonstrated that the Epstein-Barr virus-encoded latent membrane protein 1 (LMP1) potently activates the cellular c-Jun N-terminal kinase (JNK) pathway by sequentially engaging an unknown adaptor, TRAF6, TAB1/TAK1, and JNKKs. We now show that BS69, a MYND domain-containing cellular protein, is the missing adaptor that bridges LMP1 and TRAF6, as the MYND domain and a separate region of BS69 bind to the carboxyl termini of LMP1 and TRAF6, respectively. While LMP1 promotes the interaction between BS69 and TRAF6, the complex formation between LMP1 and TRAF6 is BS69 dependent. A fraction of LMP1 and BS69 is constitutively colocalized in the membrane lipid rafts. Importantly, knockdown of BS69 by small interfering RNAs specifically inhibits JNK activation by LMP1 but not tumor necrosis factor alpha. Although overexpression of either BS69 or a mutant LMP1 without the cytoplasmic carboxyl tail is not sufficient to activate JNK, interestingly, when BS69 is covalently linked to the mutant LMP1, the chimeric protein restores the ability to activate JNK. This indicates that the recruitment and aggregation of BS69 is a prerequisite for JNK activation by LMP1.

Adaptor Proteins, Vesicular Transport↗

PubMed2006

Blockade of transforming growth factor-beta-activated kinase 1 activity enhances TRAIL-induced apoptosis through activation of a caspase cascade.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL/Apo2L) is a member of the TNF-alpha ligand family that selectively induces apoptosis in a variety of tumor cells. To clarify the molecular mechanism of TRAIL-induced apoptosis, we focused on transforming growth factor-beta-activated kinase 1 (TAK1) mitogen-activated protein kinase (MAPK) kinase kinase, a key regulator of the TNF-alpha-induced activation of p65/RelA and c-Jun NH2-terminal kinase/p38 MAPKs. In human cervical carcinoma HeLa cells, TRAIL induced the delayed phosphorylation of endogenous TAK1 and its activator protein TAB1 and TAB2, which contrasted to the rapid response to TNF-alpha. Specific knockdown of TAK1 using small interfering RNA (siRNA) abrogated the TRAIL-induced activation of p65 and c-Jun NH2-terminal kinase/p38 MAPKs. TRAIL-induced apoptotic signals, including caspase-8, caspase-3, caspase-7, and poly(ADP-ribose) polymerase, were enhanced by TAK1 siRNA. Flow cytometry showed that the binding of Annexin V to cell surface was also synergistically increased by TRAIL in combination with TAK1 siRNA. In addition, pretreatment of cells with 5Z-7-oxozeaenol, a selective TAK1 kinase inhibitor, enhanced the TRAIL-induced cleavage of caspases and binding of Annexin V. The TAK1-mediated antiapoptotic effects were also observed in human lung adenocarcinoma A549 cells. In contrast, TAK1-deficient mouse embryonic fibroblasts are resistant to TRAIL-induced apoptosis, and treatment of control mouse embryonic fibroblasts with 5Z-7-oxozeaenol did not drastically promote the TRAIL-induced activation of a caspase cascade. These results suggest that TAK1 plays a critical role for TRAIL-induced apoptosis, and the blockade of TAK1 kinase will improve the chances of overcoming cancer.

Animals↗