Masanobu Nanno

The Polymeric Immunoglobulin Receptor Translocates Pneumococci across Human Nasopharyngeal Epithelial Cells

The polymeric immunoglobulin receptor (pIgR) plays a crucial role in mucosal immunity against microbial infection by transporting polymeric immunoglobulins (pIg) across the mucosal epithelium. We report here that the human pIgR (hpIgR) can bind to a major pneumococcal adhesin, CbpA. Expression of hpIgR in human nasopharyngeal cells and MDCK cells greatly enhanced pneumococcal adherence and invasion. The hpIgR-mediated bacterial adherence and invasion were abolished by either insertional knockout of cbpA or antibodies against either hpIgR or CbpA. In contrast, rabbit pIgR (rpIgR) did not bind to CbpA and its expression in MDCK cells did not enhance pneumococcal adherence and invasion. These results suggest that pneumococci are a novel example of a pathogen co-opting the pIg transcytosis machinery to promote translocation across a mucosal barrier.

Protection Against Influenza Virus Infection in Polymeric Ig Receptor Knockout Mice Immunized Intranasally with Adjuvant-Combined Vaccines

The role of secretory IgA in conferring cross-protective immunity was examined in polymeric (p)IgR knockout (KO) mice immunized intranasally with different inactivated vaccines prepared from A/PR/8/34 (H1N1), A/Yamagata/120/86 (H1N1), A/Beijing/262/95 (H1N1), and B/Ibaraki/2/85 viruses and infected with the A/PR/8/34 virus in the upper respiratory tract (RT)-restricting volume. In wild-type mice, immunization with A/PR/8/34 or its variant (A/Yamagata/120/86 and A/Beijing/262/95) vaccines conferred complete protection or partial cross-protection against infection, while the B-type virus vaccine failed to provide protection. The protection or cross-protection was accompanied by an increase in the nasal A/PR/8/34 hemagglutinin-reactive IgA concentration, which was estimated to be >30 times the serum IgA concentration and much higher than the nasal IgG concentration. In contrast, the blockade of transepithelial transport of dimeric IgA in pIgR-KO mice reduced the degree of protection or cross-protection, in parallel with the marked increase in serum IgA concentration and the decrease in nasal IgA concentration (∼20 and 0.3 times those in wild-type mice, respectively). The degree of the reduction of protection or cross-protection was moderately reversed by the low but non-negligible level of nasal IgA, transudates from the accumulated serum IgA. These results, together with the absence of the IgA-dependent cross-protection in the lower RT and the unaltered level of nasal or serum IgG in wild-type and pIgR-KO mice, confirm that the actively secreted IgA plays an important role in cross-protection against variant virus infection in the upper RT, which cannot be substituted by serum IgG.

Effects of Probiotics on Allergic Rhinitis Induced by Japanese Cedar Pollen: Randomized Double-Blind, Placebo-Controlled Clinical Trial

Background:Lactobacillus casei strain Shirota (LcS) has been found to exert antiallergic effects in animal experiments, but there is little information about its clinical effects in human patients with allergy. Methods: We performed a randomized double-blind, placebo-controlled study to investigate the effects of LcS in patients with allergic rhinitis triggered by Japanese cedar pollen (JCP). Participants were asked to drink fermented milk containing LcS (LcS group) or placebo (control group) for 8 weeks. Clinical symptoms and immunological parameters were compared between the two groups. Results: Symptom-medication scores (SMS) worsened in accordance with the increase in the amount of scattered JCP. In terms of the nasal and ocular SMS, there was no significant difference between the LcS group and the placebo group during the ingestion period. In the subgroup of patients with moderate-to-severe nasal symptom scores before starting the ingestion of test samples, supplementation with LcS tended to reduce nasal SMS. Conclusion: These results indicate that fermented milk containing LcS does not prevent allergic symptoms in patients sensitive to JCP, but may delay the occurrence of allergic symptoms in patients with moderate-to-severe nasal symptom scores.

Probiotics and immunology: separating the wheat from the chaff

Probiotics are live bacteria exhibiting health-promoting activities. Recent research has demonstrated that probiotics can prevent pathogen colonization of the gut and reduce the incidence or relieve the symptoms of various diseases caused by dysregulated immune responses. Probiotics seem to function by influencing both intestinal epithelial cells and immune cells of the gut, but the details of these effects are still being unraveled. Therefore, probiotics, through their effects on the host immune system, might ameliorate diseases triggered by disordered immune responses. Caveats remain and, because the beneficial effects of probiotics can vary between strains, the selection of the most suitable ones will be crucial for their use in the prevention or treatment of specific diseases.

γδ T cells: firefighters or fire boosters in the front lines of inflammatory responses

Summary:  Intradermal inoculation of cloned self‐reactive αβ T cells into the footpads of mice induced cutaneous graft‐versus‐host disease (GVHD), and after recovery from GVHD, the epidermis became resistant to subsequent attempts to induce GVHD. Resistance to GVHD was not induced in the epidermis of T‐cell receptor δ‐deficient (TCRδ−/−) mice that lacked γδ T cells bearing canonical Vγ5/Vδ1+γδTCRs, known as dendritic epidermal T cells (DETCs), and resistance was restored by reconstitution of these mutant mice with precursors of Vγ5+ DETCs. Pulmonary infection by Cryptococcus neoformans induced an increase of γδ T cells in the lung, and in comparison with wildtype mice, TCRδ−/− mice eliminated C. neoformans more rapidly and synthesized more interferon‐γ in the lung. In the mouse small intestine, the absence of γδ T cells is associated with a reduction in epithelial cell turnover and downregulation of the expression of major histocompatibility complex class II molecules. The protective role of γδ T cells was verified in a dextran sodium sulfate‐induced inflammatory bowel disease (IBD) model, whereas in a spontaneous model of IBD, γδ T cells were involved in the exacerbation of colitis in TCRα−/− mice. Taken together, in addition to the homeostatic regulation of epithelial tissues, γδ T cells appear to play a pivotal role in the modification of inflammatory responses induced in many organs containing epithelia.

Role of Gut Cryptopatches in Early Extrathymic Maturation of Intestinal Intraepithelial T Cells

Lympho-hemopoietic progenitors residing in murine gut cryptopatches (CP) have been shown to generate intestinal intraepithelial T cells (IEL). To investigate the role of CP in progenitor maturation, we analyzed IEL in male mice with a truncated mutation of common cytokine receptor γ-chain (CRγ−/Y) in which CP were undetectable. IEL-expressing TCR-γδ (γδ-IEL) were absent, and a drastically reduced number of Thy-1highCD4+ and Thy-1highCD8αβ+ αβ-IEL were present in CRγ−/Y mice, whereas these αβ-IEL disappeared from athymic CRγ−/Y littermate mice. Athymic CRγ−/Y mice possessed a small TCR- and αEβ7 integrin-negative IEL population, characterized by the disappearance of the extrathymic CD8αα+ subset, that expressed pre-Tα, RAG-2, and TCR-Cβ but not CD3ε transcripts. These TCR− IEL from athymic CRγ−/Y mice did not undergo Dβ-Jβ and Vδ-Jδ joinings, despite normal rearrangements at the TCR-β and -δ loci in thymocytes from euthymic CRγ−/Y mice. In contrast, athymic severe combined immunodeficient mice in which CP developed normally possessed two major TCR−αEβ7+ CD8αα+ and CD8− IEL populations that expressed pre-Tα, RAG-2, TCR-Cβ, and CD3ε transcripts. These findings underscore the role of gut CP in the early extrathymic maturation of CD8αα+ IEL, including cell-surface expression of αEβ7 integrin, CD3ε gene transcription, and TCR gene rearrangements.

Secretory IgA-mediated protection against V. cholerae and heat-labile enterotoxin-producing enterotoxigenic Escherichia coli by rice-based vaccine.

Cholera and enterotoxigenic Escherichia coli (ETEC) are among the most common causes of acute infantile gastroenteritis globally. We previously developed a rice-based vaccine that expressed cholera toxin B subunit (MucoRice-CTB) and had the advantages of being cold chain–free and providing protection against cholera toxin (CT)–induced diarrhea. To advance the development of MucoRice-CTB for human clinical application, we investigated whether the CTB-specific secretory IgA (SIgA) induced by MucoRice-CTB gives longstanding protection against diarrhea induced by Vibrio cholerae and heat-labile enterotoxin (LT)–producing ETEC (LT-ETEC) in mice. Oral immunization with MucoRice-CTB stored at room temperature for more than 3 y provided effective SIgA-mediated protection against CT- or LT-induced diarrhea, but the protection was impaired in polymeric Ig receptor–deficient mice lacking SIgA. The vaccine gave longstanding protection against CT- or LT-induced diarrhea (for ≥6 months after primary immunization), and a single booster immunization extended the duration of protective immunity by at least 4 months. Furthermore, MucoRice-CTB vaccination prevented diarrhea in the event of V. cholerae and LT-ETEC challenges. Thus, MucoRice-CTB is an effective long-term cold chain–free oral vaccine that induces CTB-specific SIgA-mediated longstanding protection against V. cholerae– or LT-ETEC–induced diarrhea.

Essential Roles of Monocytes in Stimulating Human Peripheral Blood Mononuclear Cells with Lactobacillus casei To Produce Cytokines and Augment Natural Killer Cell Activity

ABSTRACT We examined the effect of a probiotic strain, Lactobacillus casei strain Shirota, on cytokine production and natural killer (NK) cell activity in human peripheral blood mononuclear cells (PBMNC). The cellular mechanisms of immunoregulation by L. casei strain Shirota were also investigated. L. casei strain Shirota stimulated PBMNC to secrete interleukin-12 (IL-12), gamma interferon (IFN-γ), tumor necrosis factor alpha (TNF-α), and IL-10. However, depletion of monocytes from PBMNC eliminated the induction of these cytokines. L. casei strain Shirota was phagocytosed by monocytes and directly stimulated them to secrete IL-12, TNF-α, and IL-10. IFN-γ production was diminished by the addition of anti-IL-12 antibody to the PBMNC cultures. Purified T cells, but not NK cells, produced IFN-γ effectively when stimulated with L. casei strain Shirota in the presence of monocytes, indicating that monocytes triggered by L. casei strain Shirota help T cells to produce IFN-γ through secreting IL-12. In addition, NK cell activity and CD69 expression on NK cells increased after cultivation of PBMNC with L. casei strain Shirota. When monocytes were depleted from PBMNC, L. casei strain Shirota did not enhance NK cell activity. These results demonstrate that monocytes play critical roles in the induction of cytokines and following the augmentation of NK cell activity during the stimulation of human PBMNC with L. casei strain Shirota.

Curriculum vitae of intestinal intraepithelial T cells: Their developmental and behavioral characteristics

Summary:  The alimentary tract has an epithelial layer, consisting mainly of intestinal epithelial cells (IECs), that is exposed to the exterior world through the intestinal lumen. The IEC layer contains many intestinal intraepithelial T cells (IELs), and the total number of IELs constitutes the largest population in the peripheral T‐cell pool. Virtually all γδ‐IELs and many αβ‐IELs in the mouse small intestine are known to express CD8αα homodimers. A wide range of evidence that supports extrathymic development of these CD8αα+ IELs has been collected. In addition, while several studies identified cells with precursor T‐cell phenotypes within the gut epithelium, how these precursors, which are dispersed along the length of the intestine, develop into γδ‐IELs and/or αβ‐IELs has not been clarified. The identification of lymphoid cell aggregations named ‘cryptopatches’ (CPs) in the intestinal crypt lamina propria of mice as sites rich in T‐cell precursors in 1996 by our research group, however, provided evidence for a central site, whereby precursor IELs could give rise to T‐cell receptor‐bearing IELs. In this review, we discuss the development of IELs in the intestinal mucosa and examine the possibility that CPs serve as a production site of extrathymic IELs.

Peptidoglycan from lactobacilli inhibits interleukin‐12 production by macrophages induced by Lactobacillus casei through Toll‐like receptor 2‐dependent and independent mechanisms

We previously showed that Lactobacillus strains having a rigid cell wall resistant to intracellular digestion can stimulate macrophages to induce large a quantity of interleukin‐12 (IL‐12). In this study, we examined the influence of lactobacilli and bacterial cell wall components on IL‐12 production by macrophages that was induced by Lactobacillus casei, which has a rigid cell wall. Easily digestible lactobacilli such as Lactobacillus johnsonii and Lactobacillus plantarum or their intact cell walls (ICWs) weakly or very weakly induced IL‐12 production by macrophages, and inhibitedL. casei‐induced IL‐12 production. While the ICW of L. casei was resistant to intracellular digestion and did not inhibit L. casei‐induced IL‐12 production, its polysaccharide‐depleted ICW, i.e. intact peptidoglycan, was sensitive to intracellular digestion and inhibited L. casei‐induced IL‐12 production. Furthermore, the peptidoglycans of L. johnsonii, L. plantarum and Staphylococcus aureus also inhibited L. casei‐induced IL‐12 production. Peptidoglycans from lactobacilli suppressed L. casei‐induced expression of IL‐12p40 but not IL‐12p35 mRNA. Inhibition of IL‐12 production by peptidoglycan was mitigated in Toll‐like receptor 2 (TLR2)‐deficient macrophages compared with the inhibition in wild‐type macrophages. A derivative of the minimal structural unit of peptidoglycan (6‐O‐stearoyl‐muramyl dipeptide) recognized by nucleotide‐binding oligomerization domain 2 (NOD2) could also suppress L. casei‐induced IL‐12 production. These findings demonstrate that easily digestible bacteria and peptidoglycan suppress IL‐12 production through pattern recognition receptors such as TLR2 and NOD2. IL‐12 production in the gut may be negatively regulated by the simultaneous inhibitory actions of various resident bacteria that are susceptible to intracellular digestion.