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Jun Kunisawa,
Masashi Gohda,
Eri Hashimoto,
Izumi Ishikawa,
Morio Higuchi,
Yuji Suzuki,
Yoshiyuki Goto,
Casandra Panea,
Ivaylo I Ivanov,
Risa Sumiya,
Lamichhane Aayam,
Taichi Wake,
So Tajiri,
Yosuke Kurashima,
Shiori Shikata,
Shizuo Akira,
Kiyoshi Takeda, Hiroshi Kiyono
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ABSTRACT: Intestinal plasma cells predominantly produce immunoglobulin (Ig) A, however, their functional diversity remains poorly characterized. Here we show that murine intestinal IgA plasma cells can be newly classified into two populations on the basis of CD11b expression, which cannot be discriminated by currently known criteria such as general plasma cell markers, B cell origin and T cell dependence. CD11b(+) IgA(+) plasma cells require the lymphoid structure of Peyer's patches, produce more IgA than CD11b(-) IgA(+) plasma cells, proliferate vigorously, and require microbial stimulation and IL-10 for their development and maintenance. These features allow CD11b(+) IgA(+) plasma cells to mediate early-phase antigen-specific intestinal IgA responses induced by oral immunization with protein antigen. These findings reveal the functional diversity of IgA(+) plasma cells in the murine intestine.
Nature Communications 04/2013; 4:1772. · 7.40 Impact Factor
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Yoshikazu Yuki,
Mio Mejima,
Shiho Kurokawa,
Tomoko Hiroiwa,
Yuko Takahashi,
Daisuke Tokuhara,
Tomonori Nochi,
Yuko Katakai,
Masaharu Kuroda,
Natsumi Takeyama, [......],
Michiyo Abe,
Yingju Chen,
Ushio Nakanishi,
Takehiro Masumura,
Yoji Takeuchi,
Hiroko Kozuka-Hata,
Hiroaki Shibata,
Masaaki Oyama,
Kunisuke Tanaka, Hiroshi Kiyono
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ABSTRACT: Plants have been used as expression systems for a number of vaccines. However, the expression of vaccines in plants sometimes results in unexpected modification of the vaccines by N-terminal blocking and sugar-chain attachment. Although MucoRice-CTB was thought to be the first cold-chain-free and unpurified oral vaccine, the molecular heterogeneity of MucoRice-CTB, together with plant-based sugar modifications of the CTB protein, has made it difficult to assess immunological activity of vaccine and yield from rice seed. Using a T-DNA vector driven by a prolamin promoter and a signal peptide added to an overexpression vaccine cassette, we established MucoRice-CTB/Q as a new generation oral cholera vaccine for humans use. We confirmed that MucoRice-CTB/Q produces a single CTB monomer with an Asn to Gln substitution at the 4th glycosylation position. The complete amino acid sequence of MucoRice-CTB/Q was determined by MS/MS analysis and the exact amount of expressed CTB was determined by SDS-PAGE densitometric analysis to be an average of 2.35 mg of CTB/g of seed. To compare the immunogenicity of MucoRice-CTB/Q, which has no plant-based glycosylation modifications, with that of the original MucoRice-CTB/N, which is modified with a plant N-glycan, we orally immunized mice and macaques with the two preparations. Similar levels of CTB-specific systemic IgG and mucosal IgA antibodies with toxin-neutralizing activity were induced in mice and macaques orally immunized with MucoRice-CTB/Q or MucoRice-CTB/N. These results show that the molecular uniformed MucoRice-CTB/Q vaccine without plant N-glycan has potential as a safe and efficacious oral vaccine candidate for human use.
Plant Biotechnology Journal 04/2013; · 5.44 Impact Factor
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Ilgyu Kong,
Ayuko Sato,
Yoshikazu Yuki,
Tomonori Nochi,
Haruko Takahashi,
Shinichi Sawada,
Mio Mejima,
Shiho Kurokawa,
Kazunari Okada,
Shintaro Sato,
David E Briles,
Jun Kunisawa,
Yusuke Inoue,
Masafumi Yamamoto,
Kazunari Akiyoshi, Hiroshi Kiyono
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ABSTRACT: To establish a safer and more effective vaccine against pneumococcal respiratory infections, current knowledge regarding the antigens common among pneumococcal strains and improvements to the system for delivering these antigens across the mucosal barrier must be integrated. We developed a pneumococcal vaccine that combines the advantages of pneumococcal surface protein A (PspA) with a nontoxic intranasal vaccine-delivery system based on a nanometer-sized hydrogel (nanogel) bearing a cationic cholesteryl-group-bearing pullulan (cCHP). The efficacy of the nanogel-based PspA nasal vaccine (cCHP-PspA) was tested in murine pneumococcal airway infection models. Intranasal vaccination with cCHP-PspA provided protective immunity against lethal challenge with Streptococcus pneumoniae Xen10, reduced colonization and invasion by bacteria in the upper and lower respiratory tracts, and induced systemic and nasal mucosal Th17 responses, high levels of PspA-specific serum IgG, and nasal and bronchial IgA antibody responses. Moreover, there was no sign of PspA delivery by nanogel to either the olfactory bulbs or the central nervous system after intranasal administration. These results demonstrate the effectiveness and safety of the nanogel-based PspA nasal vaccine system as a universal mucosal vaccine against pneumococcal respiratory infection.
Infection and immunity 03/2013; · 4.21 Impact Factor
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Yoshiko Fukuyama,
Daisuke Tokuhara,
Shinichi Sekine,
Kazuyoshi Aso,
Kosuke Kataoka,
Julia Davydova,
Masato Yamamoto,
Rebekah S Gilbert,
Yuka Tokuhara,
Keiko Fujihashi,
Jun Kunisawa,
Yoshikazu Yuki, Hiroshi Kiyono,
Jerry R McGhee,
Kohtaro Fujihashi
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ABSTRACT: We assessed the role of CCR5(+)/CCR6(+)/CD11b(+)/CD11c(+) dendritic cells (DCs) for induction of ovalbumin (OVA)-specific antibody (Ab) responses following mucosal immunization. Mice given nasal OVA plus an adenovirus expressing Flt3 ligand (Ad-FL) showed early expansion of CCR5(+)/CCR6(+)/CD11b(+)/CD11c(+) DCs in nasopharyngeal-associated lymphoid tissue (NALT) and cervical lymph nodes (CLNs). Subsequently, this DC subset became resident in submandibular glands (SMGs) and nasal passages (NPs) in response to high levels of CCR-ligands produced in these tissues. CD11b(+)/CD11c(+) DCs were markedly decreased in both CCR5(-/-) and CCR6(-/-) mice. Chimera mice reconstituted with bone marrow cells from CD11c-diphtheria toxin receptor (CD11c-DTR) and CCR5(-/-) or CD11c-DTR and CCR6(-/-) mice given nasal OVA plus Ad-FL had elevated plasma IgG, but reduced IgA as well as low anti-OVA secretory IgA (SIgA )Ab responses in saliva and nasal washes. These results suggest that CCR5(+)CCR6(+) DCs play an important role in the induction of Ag-specific SIgA Ab responses.
PLoS ONE 01/2013; 8(4):e60453. · 4.09 Impact Factor
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Yosuke Kurashima,
Takeaki Amiya,
Tomonori Nochi,
Kumiko Fujisawa,
Takeshi Haraguchi,
Hideo Iba,
Hiroko Tsutsui,
Shintaro Sato,
Sachiko Nakajima,
Hideki Iijima,
Masato Kubo,
Jun Kunisawa, Hiroshi Kiyono
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ABSTRACT: Mast cells are known effector cells in allergic and inflammatory diseases, but their precise roles in intestinal inflammation remain unknown. Here we show that activation of mast cells in intestinal inflammation is mediated by ATP-reactive P2X7 purinoceptors. We find an increase in the numbers of mast cells expressing P2X7 purinoceptors in the colons of mice with colitis and of patients with Crohn's disease. Treatment of mice with a P2X7 purinoceptor-specific antibody inhibits mast cell activation and subsequent intestinal inflammation. Similarly, intestinal inflammation is ameliorated in mast cell-deficient Kit(W-sh/W-sh) mice, and reconstitution with wild-type, but not P2x7(-/-) mast cells results in susceptibility to inflammation. ATP-P2X7 purinoceptor-mediated activation of mast cells not only induces inflammatory cytokines, but also chemokines and leukotrienes, to recruit neutrophils and subsequently exacerbate intestinal inflammation. These findings reveal the role of P2X7 purinoceptor-mediated mast cell activation in both the initiation and exacerbation of intestinal inflammation.
Nature Communications 09/2012; 3:1034. · 7.40 Impact Factor
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ABSTRACT: The intestinal enterocytes and other epithelial cells create physical barriers, including tight junctions and mucus layers. These cells also actively transport antibodies across the epithelium and simultaneously produce antimicrobial peptides and enzymes. These functions maintain intestinal homeostasis by allowing the selective absorption of nutrients and simultaneously preventing pathogenic infections. Recent evidence has revealed that both host-derived factors (e.g., cytokines) and gut environmental factors (e.g., commensal bacteria, dietary materials, and their metabolites) regulate the physical and immunological functions of the epithelium. Understanding the interactions between host cells and these environmental factors should help us to develop new strategies to prevent and treat immune diseases of the intestine.
Drug discovery today 08/2012; · 6.63 Impact Factor
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Takuma Shibata,
Naoki Takemura,
Yuji Motoi,
Yoshiyuki Goto,
Thangaraj Karuppuchamy,
Kumi Izawa,
Xiaobing Li,
Sachiko Akashi-Takamura,
Natsuko Tanimura,
Jun Kunisawa, Hiroshi Kiyono,
Shizuo Akira,
Toshio Kitamura,
Jiro Kitaura,
Satoshi Uematsu,
Kensuke Miyake
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ABSTRACT: 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.
International Immunology 07/2012; 24(10):613-23. · 3.41 Impact Factor
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Yoshikazu Yuki,
Mio Mejima,
Shiho Kurokawa,
Tomoko Hiroiwa,
Il Gyu Kong,
Masaharu Kuroda,
Yoko Takahashi,
Tomonori Nochi,
Daisuke Tokuhara,
Tomoko Kohda,
Shunji Kozaki, Hiroshi Kiyono
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ABSTRACT: Mucosal vaccines based on rice (MucoRice) offer a highly practical and cost-effective strategy for vaccinating large populations against mucosal infections. However, the limitation of low expression and yield of vaccine antigens with high molecular weight remains to be overcome. Here, we introduced RNAi technology to advance the MucoRice system by co-introducing antisense sequences specific for genes encoding endogenous rice storage proteins to minimize storage protein production and allow more space for the accumulation of vaccine antigen in rice seed. When we used RNAi suppression of a combination of major rice endogenous storage proteins, 13 kDa prolamin and glutelin A in a T-DNA vector, we could highly express a vaccine comprising the 45 kDa C-terminal half of the heavy chain of botulinum type A neurotoxin (BoHc), at an average of 100 μg per seed (MucoRice-BoHc). The MucoRice-Hc was water soluble, and was expressed in the cytoplasm but not in protein body I or II of rice seeds. Thus, our adaptation of the RNAi system improved the yield of a vaccine antigen with a high molecular weight. When the mucosal immunogenicity of the purified MucoRice-BoHc was examined, the vaccine induced protective immunity against a challenge with botulinum type A neurotoxin in mice. These findings demonstrate the efficiency and utility of the advanced MucoRice system as an innovative vaccine production system for generating highly immunogenic mucosal vaccines of high-molecular-weight antigens.
Vaccine 04/2012; 30(28):4160-6. · 3.77 Impact Factor
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ABSTRACT: Most viruses use host mucosal surfaces as their initial portals of infection. The respiratory tract has the body's second-largest mucosal surface area after the digestive tract. An understanding of the unique nature of the mucosal immune system of respiratory organs is therefore extremely important for the development of new-generation vaccines and novel methods of preventing and treating respiratory infectious diseases, including viral infections.
Current opinion in virology. 04/2012; 2(3):225-32.
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ABSTRACT: To develop protective immune responses against mucosal pathogens, the delivery route and adjuvants for vaccination are important. The host, however, strives to maintain mucosal homeostasis by responding to mucosal antigens with tolerance, instead of immune activation. Thus, induction of mucosal immunity through vaccination is a rather difficult task, and potent mucosal adjuvants, vectors or other special delivery systems are often used, especially in the elderly. By taking advantage of the common mucosal immune system, the targeting of mucosal dendritic cells and microfold epithelial cells may facilitate the induction of effective mucosal immunity. Thus, novel routes of immunization and antigen delivery systems also show great potential for the development of effective and safe mucosal vaccines against various pathogens. The purpose of this review is to introduce several recent approaches to induce mucosal immunity to vaccines, with an emphasis on mucosal tissue targeting, new immunization routes and delivery systems. Defining the mechanisms of mucosal vaccines is as important as their efficacy and safety, and in this article, examples of recent approaches, which will likely accelerate progress in mucosal vaccine development, are discussed.
Expert Review of Vaccines 03/2012; 11(3):367-79. · 4.25 Impact Factor
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ABSTRACT: It has been shown that dietary materials are involved in immune regulation in the intestine. Lipids mediate immune regulation through a complex metabolic network that produces many kinds of lipid mediators. Sphingosine-1-phosphate (S1P) is a lipid mediator that controls cell trafficking and activation. In this review, we focus on the immunological functions of S1P in the regulation of intestinal immune responses such as immunoglobulin A production and unique T cell trafficking, and its role in the development of intestinal immune diseases such as food allergies and intestinal inflammation, and also discuss the relationship between dietary materials and S1P metabolism.
Nutrients 03/2012; 4(3):154-66. · 0.68 Impact Factor
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ABSTRACT: Secretory-immunoglobulin A (S-IgA) plays an important role in immunological defense in the intestine. It has been known for a long time that microbial stimulation is required for the development and maintenance of intestinal IgA production. Recent advances in genomic technology have made it possible to detect uncultivable commensal bacteria in the intestine and identify key bacteria in the regulation of innate and acquired mucosal immune responses. In this review, we focus on the immunological function of Peyer's patches (PPs), a major gut-associated lymphoid tissue, in the induction of intestinal IgA responses and the unique immunological interaction of PPs with commensal bacteria, especially Alcaligenes, a unique indigenous bacteria habituating inside PPs.
Frontiers in immunology. 01/2012; 3:65.
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ABSTRACT: Large numbers of environmental antigens, including commensal bacteria and food-derived antigens, constitutively interact with the epithelial layer of the gastrointestinal (GI) tract. Commensal bacteria peacefully cohabit with the host GI tract and exert multiple beneficial or destructive effects on their host. Intestinal epithelial cells (IECs) constitute the first physical and immunological protective wall against invasive pathogens and a cohabitation niche for commensal bacteria. As the physiological homeostasis of IECs is maintained by multiple biological processes such as apoptosis, autophagy, and the handling of endoplasmic reticulum stress, the aberrant kinetics of these biological events, which have genetic and environmental causes, leads to the development of host intestinal pathogenesis such as inflammatory bowel disease. In addition, IECs recognize and interact with commensal bacteria and give instructions to mucosal immune cells to initiate an immunological balance between active and quiescent conditions, eventually establishing intestinal homeostasis. The mucosal immune system regulates the homeostasis of gut microbiota by producing immunological molecules such as secretory immunoglobulin A, the production of which is mediated by IECs. IECs therefore play a central role in the creation and maintenance of a physiologically and immunologically stable intestinal environment.
Immunological Reviews 01/2012; 245(1):147-63. · 11.15 Impact Factor
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ABSTRACT: Dietary factors regulate immunological function, but the underlying mechanisms remain elusive. Here we show that vitamin B9 is a survival factor for regulatory T (Treg) cells expressing high levels of vitamin B9 receptor (folate receptor 4). In vitamin B9-reduced condition in vitro, Treg cells could be differentiated from naïve T cells but failed to survive. The impaired survival of Treg cells was associated with decreased expression of anti-apoptotic Bcl2 and independent of IL-2. In vivo depletion of dietary vitamin B9 resulted in the reduction of Treg cells in the small intestine, a site for the absorption of dietary vitamin B9. These findings provide a new link between diet and the immune system, which could maintain the immunological homeostasis in the intestine.
PLoS ONE 01/2012; 7(2):e32094. · 4.09 Impact Factor
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ABSTRACT: Mucosal vaccination has several advantages compared with that of injection-type vaccination. Secretory IgA(SIgA) produced at mucosal surface plays a key role for inactivation of toxins and inhibition of pathogen invasion. Although oral or nasal vaccination with attenuated live microorganisms have been shown to be effective in the induction of protective immunity, these types of vaccine have the ability to infect transiently to the host. For the development of safe and effective mucosal vaccine, an obvious strategy is the preparation of inactivated subunit-type mucosal vaccine. Here we introduce our frontier technology for the development of rice-based oral vaccines, as a new generation of mucosal vaccine. Further, we also discuss recent progress in the development of other types of mucosal vaccine and adjuvant.
Nippon rinsho. Japanese journal of clinical medicine 09/2011; 69(9):1555-60.
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ABSTRACT: Oral vaccine has been considered to be a prospective vaccine against many pathogens especially invading across gastrointestinal tracts. One key element of oral vaccine is targeting efficient delivery of antigen to gut-associated lymphoid tissue (GALT), the inductive site in the intestine where antigen-specific immune responses are initiated. Various chemical and biological antigen delivery systems have been developed and some are in clinical trials. In this review, we describe the immunological features of GALT and the current status of antigen delivery system candidates for successful oral vaccine.
Advanced drug delivery reviews 07/2011; 64(6):523-30. · 11.96 Impact Factor
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Michinaga Ogawa,
Yuko Yoshikawa,
Taira Kobayashi,
Hitomi Mimuro,
Makoto Fukumatsu,
Kotaro Kiga,
Zhenzi Piao,
Hiroshi Ashida,
Mitsutaka Yoshida,
Shigeru Kakuta,
Tomohiro Koyama,
Yoshiyuki Goto,
Takahiro Nagatake,
Shinya Nagai, Hiroshi Kiyono,
Magdalena Kawalec,
Jean-Marc Reichhart,
Chihiro Sasakawa
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ABSTRACT: Selective autophagy of bacterial pathogens represents a host innate immune mechanism. Selective autophagy has been characterized on the basis of distinct cargo receptors but the mechanisms by which different cargo receptors are targeted for autophagic degradation remain unclear. In this study we identified a highly conserved Tectonin domain-containing protein, Tecpr1, as an Atg5 binding partner that colocalized with Atg5 at Shigella-containing phagophores. Tecpr1 activity is necessary for efficient autophagic targeting of bacteria, but has no effect on rapamycin- or starvation-induced canonical autophagy. Tecpr1 interacts with WIPI-2, a yeast Atg18 homolog and PI(3)P-interacting protein required for phagophore formation, and they colocalize to phagophores. Although Tecpr1-deficient mice appear normal, Tecpr1-deficient MEFs were defective for selective autophagy and supported increased intracellular multiplication of Shigella. Further, depolarized mitochondria and misfolded protein aggregates accumulated in the Tecpr1-knockout MEFs. Thus, we identify a Tecpr1-dependent pathway as important in targeting bacterial pathogens for selective autophagy.
Cell host & microbe 05/2011; 9(5):376-89. · 13.02 Impact Factor
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Nature Immunology 03/2011; 12(3):195-7. · 26.01 Impact Factor
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ABSTRACT: Quantitative and qualitative aspects of commensal bacteria determine the active and quiescent status of host immunity. In a recent Science paper, Atarashi et al. (2011) identify Clostridium clusters IV and XIVa as indigenous commensal bacteria that induce regulatory T cells for the creation and maintenance of immunological homeostasis.
Cell host & microbe 02/2011; 9(2):83-4. · 13.02 Impact Factor
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Dong-Young Kim,
Ayuko Sato,
Satoshi Fukuyama,
Hiroshi Sagara,
Takahiro Nagatake,
Il Gyu Kong,
Kaoru Goda,
Tomonori Nochi,
Jun Kunisawa,
Shintaro Sato,
Yoshifumi Yokota,
Chul Hee Lee, Hiroshi Kiyono
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ABSTRACT: In this study, we demonstrated a new airway Ag sampling site by analyzing tissue sections of the murine nasal passages. We revealed the presence of respiratory M cells, which had the ability to take up OVA and recombinant Salmonella typhimurium expressing GFP, in the turbinates covered with single-layer epithelium. These M cells were also capable of taking up respiratory pathogen group A Streptococcus after nasal challenge. Inhibitor of DNA binding/differentiation 2 (Id2)-deficient mice, which are deficient in lymphoid tissues, including nasopharynx-associated lymphoid tissue, had a similar frequency of M cell clusters in their nasal epithelia to that of their littermates, Id2(+/-) mice. The titers of Ag-specific Abs were as high in Id2(-/-) mice as in Id2(+/-) mice after nasal immunization with recombinant Salmonella-ToxC or group A Streptococcus, indicating that respiratory M cells were capable of sampling inhaled bacterial Ag to initiate an Ag-specific immune response. Taken together, these findings suggest that respiratory M cells act as a nasopharynx-associated lymphoid tissue-independent alternative gateway for Ag sampling and subsequent induction of Ag-specific immune responses in the upper respiratory tract.
The Journal of Immunology 02/2011; 186(7):4253-62. · 5.79 Impact Factor
