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New animal model of shigellosis in the guinea pig: Its usefulness for protective efficacy studies |
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It has been difficult to evaluate the protective efficacy of vaccine candidates against shigellosis, a major form of bacillary dysentery caused by Shigella spp. infection, because of the lack of suitable animal models. To develop a proper animal model representing human bacillary dysentery, guinea pigs were challenged with virulent Shigella flexneri setotype 2a (strains 2457T or YSH6000) or S. flexneri 5a (strains M90T) by the intrarectal (i.r.) route. Interestingly, all guinea pigs administered these Shigella strains developed severe and acute rectocolitis. They lost ~20% of their body weight and developed tenesmus by 24 h after Shigella infection. Shigella invasion and colonization of the distal colon were seen at 24 h but disappeared by 48 h following i.r. infection. Histopathological approaches demonstrated significant damage and destruction of mucosal and submucosal layers, thickened intestinal wall, edema, erosion, infiltration of neutrophils, and depletion of goblet cells in the distal colon. Furthermore, robust expression of IL-8, IL-1-¥â; and inducible NO synthase mRNA was detected in the colon from 6 h to 24 h following Shigella infection. Most importantly, in our new shigellosis model, guinea pigs vaccinated with an attenuated S. flexneri 2a SC602 strain possessing high levels of mucosal IgA Abs showed milder symptoms of bacillary dysentery than did those receiving PBS alone after Shigella infection. In the guinea pig, administration of Shigella by the i.r. route induces acute inflammation, making this a useful animal model for assessing the protective efficacy of Shigella vaccine candidates. |
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| 2. |
Sublingual mucosa is a potent route for targeting influenza virus infection
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Recent outbreaks of highly pathogenic avian influenza virus in poultry and in humans have brought home as never before the need for an effective vaccine delivery system for the prevention of influenza pandemics. We attempted to determine if the sublingual (s.l.) route would be an effective means of delivering vaccine against influenza virus by immunizing mice twice via the s.l. route with either formalin-inactivated influenza A/PR8 virus (H1N1) plus mucosal adjuvants (native CT or mutant CTA/LTB) or with a single dose of live A/PR8 virus. Interestingly, s.l. immunization resulted in both systemic and mucosal Ig responses and in strong protective efficacy against a lethal dose of influenza virus challenging via intranasal (i.n.) route. In addition, s.l. administration with formalin-inactivated A/PR8 virus plus mucosal adjuvants prompted CD8+ T cells to secrete more IFN-???resulting in dominant virus-specific cytotoxic T lymphocyte responses. Further, s.l. administration enhanced hetero-subtypic immunity, defined as protective cross-reactivity to different serotypes of virus. Most interestingly, in contrast to the i.n. route, the A/PR8 virus, whether in its live or inactivated form, did not migrate to and replicate in the central nervous system after s.l. administration. Based upon these findings, we propose here that the s.l. mucosa is a highly promising and attractive alternative to current mucosal vaccine delivery sites for targeting influenza virus infection. |
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Langerhans cell trafficking to the mesenteric lymph node sets the stage for skin and gut immune system cross-talk |
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As a topical needle-free vaccine delivery method, transcutaneous immunization (TCI) presents many clinical advantages, but its underlying mechanism remains unknown. TCI of mice with tetanus toxoid and cholera toxin induced Ag-specific IgA Ab-secreting cells (ASCs) in the small intestine in addition to IgG Abs in the sera. The IgA ASCs in the intestine which expressed functional CCR9 and CCR10 were maintained in the Peyer¡¯s patch (PP)-null mice but abolished in the PP- and LN- null mice after TCI. Moreover, the number of Ag-specific CD4+ T cells expressing ?4?7 integrins and CCR9 was increased in the MLN but not in the PP. The MLN was shown to be the site of IgA isotype class switching recombination against Ag delivered from the skin. Strikingly, an emergent population of Langerin+CD205+ cells appeared in the MLN after TCI. TCI did not affect serum Ab levels but significantly decreased intestinal IgA ASCs in both Langerhans cells (LCs)-depleted and CCR7-/- mice implying that trafficking of skin LCs into the MLN could be involved in the induction of intestinal IgA Ab responses after skin immunization. Taken together, MLN then seems to be indispensable for the induction of intestinal SIgA Abs following TCI. These results suggest a new paradigm for cross-talk between the skin and gut immune systems by LCs via the MLN. |
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| 4. |
Colonic patches direct the cross-talk between systemic compartments and the large intestine independently of innate immunity |
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Although the mucosal immune and the systemic immune systems are strucutrually and functionally independent, they do seem to engage in cross-talk under specific conditions. To further investigate this cross-talk, we vaccinated mice with tetanus toxoid together with cholera toxin as mucosal adjuvant first by the subcutaneous (SC) and then by the intrarectal (IR) route (SC-IR). Interestingly, higher numbers of Ag-specific IgA and IgG Ab-secreting cells (ASC) were detected in the lamina propria (LP) of the large intestine (LI) of SC-IR-vaccinated than in that of mice vaccinated by SC or IR alone. In SC-IR-vaccinated mice, Ag-specific ASCs from LI-LP migrated only after treatment with SDF-1?/CXCR4 and MEC/CCR10, while MEC/CCR10 alone was required for such migration in mice receiving two IR administrations. These findings suggest that Ag-specific CXCR4- and CCR10-expressing IgA ASCs detected in the LI after SC-IR might originate from systemically committed cells. After SC-IR, Ag-specific ASCs in the LI were depleted in mice lacking colonic patches (CP), suggesting that CP might be a major route for Ag delivery to recruit systemic-derived cells. Furthermore, the higher expression of IgA class switch recombination-related molecules and the accumulation of IgA ASCs when treated with FTY720 in the CP revealed that CP is the IgA class switching site after SC-IR. Interestingly, the combined SC-IR vaccination resulted in numbers of Ag-specific IgA ASCs in the LI of TLR2-/-, TLR4-/- and MyD88-/- mice that were comparable with those of wild-type mice. These results suggest that innate immunity may not direct the cross-talk communication between LI and systemic compartments following SC-IR. Taken together, our results suggest the possibility that cross-talk could occur between LI and systemic immune systems via the CP and without the assistance of innate immunity. |
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| 5. |
Enhancement of mucosal IgA Ab production following oral vaccination with attenuated Salmonella expressing PspA in the absence of MyD88 signaling |
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Toll-like receptors (TLRs) detect microbial signature molecules and play a crucial role in the induction of immune responses. TLRs are known to directly induce innate host defense responses, but the mechanisms of TLR-mediated adaptive immunity remain subject to debate. Provided that there is TLR signaling, dendritic cells mature in response to microbial infection in vitro; however, the role of such signaling in B cell activation of mucosal immunity in vivo is poorly understood. Here we show that Ab responses at mucosal sites are controlled by TLR signaling following oral administration with recombinant attenuated Salmonella (S.) typhimurium vaccine (RASV) strains expressing pneumonia surface protein A (PspA). Interestingly, oral vaccination with RASV expressing PspA resulted in significantly higher levels of PspA-specific IgA Ab responses in the fecal extracts of MyD88-/- mice than in those of wild-type, TLR4-/-, and TLR5-/- mice. Similarly, higher levels of PspA-specific IgA Ab-secreting cells were detected in the small and large intestines of MyD88-/- mice than of wild-type mice. The ratio of IgG2a (Th1) to IgG1 (Th2) Ab in the serum decreased significantly more in the MyD88-/- mice (0.84) than in the wild-type mice (1.33), indicating a Th2-dominant condition in the MyD88-/- mice following oral vaccination with RASV expressing PspA. Taken together, these findings suggest that mucosal IgA responses are accelerated in the absence of MyD88 signaling following oral microbial challenge, perhaps due to the Th2 environment that predominates in these mice. |
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