BackgroundVitamin D3, the most physiologically relevant form of vitamin D, is an essential organic compound that has been shown to have a crucial effect on the immune responses. Vitamin D3 ameliorates the onset of the experimental autoimmune encephalomyelitis (EAE); however, the direct effect of vitamin D3 on T cells is largely unknown.Methodology/Principal FindingsIn an in vitro system using cells from mice, the active form of vitamin D3 (1,25-dihydroxyvitamin D3) suppresses both interleukin (IL)-17-producing T cells (TH17) and regulatory T cells (Treg) differentiation via a vitamin D receptor signal. The ability of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) to reduce the amount of IL-2 regulates the generation of Treg cells, but not TH17 cells. Under TH17-polarizing conditions, 1,25(OH)2D3 helps to increase the numbers of IL-10-producing T cells, but 1,25(OH)2D3's negative regulation of TH17 development is still defined in the IL-10−/− T cells. Although the STAT1 signal reciprocally affects the secretion of IL-10 and IL-17, 1,25(OH)2D3 inhibits IL-17 production in STAT1−/− T cells. Most interestingly, 1,25(OH)2D3 negatively regulates CCR6 expression which might be essential for TH17 cells to enter the central nervous system and initiate EAE.Conclusions/SignificanceOur present results in an experimental murine model suggest that 1,25(OH)2D3 can directly regulate T cell differentiation and could be applied in preventive and therapeutic strategies for TH17-mediated autoimmune diseases.

Retinoic acid (RA), a well-known vitamin A metabolite, mediates inhibition of the IL-6-driven induction of proinflammatory Th17 cells and promotes anti-inflammatory regulatory T cell generation in the presence of TGF-β, which is mainly regulated by dendritic cells. To directly address the role of RA in Th17/regulatory T cell generation in vivo, we generated vitamin A-deficient (VAD) mice by continuous feeding of a VAD diet beginning in gestation. We found that a VAD diet resulted in significant inhibition of Th17 cell differentiation in the small intestine lamina propria by as early as age 5 wk. Furthermore, this diet resulted in low mRNA expression levels of IL-17, IFN regulatory factor 4, IL-21, IL-22, and IL-23 without alteration of other genes, such as RORγt, TGF-β, IL-6, IL-25, and IL-27 in the small intestine ileum. In vitro results of enhanced Th17 induction by VAD dendritic cells did not mirror in vivo results, suggesting the existence of other regulation factors. Interestingly, the VAD diet elicited high levels of mucin MUC2 by goblet cell hyperplasia and subsequently reduced gut microbiome, including segmented filamentous bacteria. Much like wild-type mice, the VAD diet-fed MyD88−/−TRIF−/− mice had significantly fewer IL-17–secreting CD4+ T cells than the control diet-fed MyD88−/−TRIF−/− mice. The results strongly suggest that RA deficiency altered gut microbiome, which in turn inhibited Th17 differentiation in the small intestine lamina propria.

We have recently reported that the sublingual (s.l.) mucosa is an efficient site for inducing systemic and mucosal immune responses. In this study, the potential of s.l. immunization to induce remote Ab responses and CD8+ cytotoxic responses in the female genital tract was examined in mice by using a nonreplicating Ag, OVA, and cholera toxin (CT) as an adjuvant. Sublingual administration of OVA and CT induced Ag-specific IgA and IgG Abs in blood and in cervicovaginal secretions. These responses were associated with large numbers of IgA Ab-secreting cells (ASCs) in the genital mucosa. Genital ASC responses were similar in magnitude and isotype distribution after s.l., intranasal, or vaginal immunization and were superior to those seen after intragastric immunization. Genital, but not blood or spleen, IgA ASC responses were inhibited by treatment with anti-CCL28 Abs, suggesting that the chemokine CCL28 plays a major role in the migration of IgA ASC progenitors to the reproductive tract mucosa. Furthermore, s.l. immunization with OVA induced OVA-specific effector CD8+ cytolytic T cells in the genital mucosa, and these responses required coadministration of the CT adjuvant. Furthermore, s.l. administration of human papillomavirus virus-like particles with or without the CT adjuvant conferred protection against genital challenge with human papillomavirus pseudovirions. Taken together, these findings underscore the potential of s.l. immunization as an efficient vaccination strategy for inducing genital immune responses and should impact on the development of vaccines against sexually transmitted diseases.

The external part of the eye shares mucosa-associated common characteristics and is an obvious entry site for foreign Ags. We assessed the potential of eyedrop vaccination for effective delivery of vaccines against viral or bacterial infection in mice. Both OVA-specific IgG Ab in serum and IgA Ab in mucosal compartments were induced by eyedrops of OVA with cholera toxin (CT). Eyedrop vaccination of influenza A/PR/8 virus (H1N1) induced both influenza virus-specific systemic and mucosal Ab responses and protected mice completely against respiratory infection with influenza A/PR/8 virus. In addition, eyedrop vaccination of attenuated Salmonella vaccine strains induced LPS-specific Ab and complete protection against oral challenge of virulent Salmonella. Unlike with the intranasal route, eyedrop vaccinations did not redirect administered Ag into the CNS in the presence of CT. When mice were vaccinated by eyedrop, even after the occlusion of tear drainage from eye to nose, Ag-specific systemic IgG and mucosal IgA Abs could be induced effectively. Of note, eyedrops with OVA plus CT induced organogenesis of conjunctiva-associated lymphoid tissue and increased microfold cell-like cells on the conjunctiva-associated lymphoid tissue in the nictitating membrane on conjunctiva, the mucosal side of the external eye. On the basis of these findings, we propose that the eyedrop route is an alternative to mucosal routes for administering vaccines.

Topical transcutaneous immunization (TCI) presents many clinical advantages, but its underlying mechanism remains unknown. TCI induced Ag-specific IgA Ab-secreting cells expressing CCR9 and CCR10 in the small intestine in a retinoic acid-dependent manner. These intestinal IgA Abs were maintained in Peyer’s patch-null mice but abolished in the Peyer’s patch- and lymph node-null mice. The mesenteric lymph node (MLN) was shown to be the site of IgA isotype class switching after TCI. Unexpectedly, langerin+CD8α− dendritic cells emerged in the MLN after TCI; they did not migrate from the skin but rather differentiated rapidly from bone marrow precursors. Depletion of langerin+ cells impaired intestinal IgA Ab responses after TCI. Taken together, these findings suggest that MLN is indispensable for the induction of intestinal IgA Abs following skin immunization and that cross-talk between the skin and gut immune systems might be mediated by langerin+ dendritic cells in the MLN.

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Previous studies demonstrated cross talk between mucosal and reproductive organs during secretory IgA Ab induction. In this study, we aimed to clarify the underlying mechanisms of this cross talk. We found significantly higher titers of Ag-specific secretory IgA Ab in the vaginal wash after mucosal vaccination by both the intranasal (i.n.) and the intravaginal routes but not by the s.c. route. Interestingly, Ag-specific IgA Ab-secreting cells (ASCs) were found mainly in the uterus but not in the cervix and vaginal canal after i.n. vaccination. The fact that most Ag-specific IgA ASCs isolated from the uteri of vaccinated mice migrated toward mucosa-associated epithelial chemokine (MEC)/CCL28 suggests dominant expression of CCR10 on the IgA ASCs. Further, IgA ASCs in the uteri of vaccinated mice were reduced drastically in mice treated with neutralizing anti-MEC/CCL28 Ab. Most intriguingly, the female sex hormone estrogen directly regulated MEC/CCL28 expression and was augmented by i.n. vaccination with cholera toxin or stimulators for innate immunity. Further, blockage of estrogen function in the uterus by oral administration of the estrogen antagonist raloxifene significantly inhibited migration of Ag-specific IgA ASCs after i.n. vaccination with OVA plus cholera toxin. Taken together, these data strongly suggest that CCR10+ IgA ASCs induced by mucosal vaccination via the i.n. route migrate into the uterus in a MEC/CCL28-dependent manner and that estrogen might have a crucial role in the protection against genital infection by regulating MEC/CCL28 expression in the uterus.

Background: Chitinase 3-like 1 protein (CHI3L1) (YKL-40 in humans and breast regression protein [BRP]-39 in mice) is required for optimal allergen sensitization and Th2 inflammation in various chronic inflammatory diseases including asthma. However, the role of CHI3L1 in airway inflammation induced by respiratory viruses has not been investigated. The aim of this study was to investigate the relationship between CHI3L1 and airway inflammation caused by respiratory syncytial virus (RSV) infection. Methods:We measured YKL-40 levels in human nasopharyngeal aspirate (NPA) from hospitalized children presenting with acute respiratory symptoms. Wild-type (WT) and BRP-39 knockout (KO) C57BL/6 mice were inoculated with live RSV (A2 strain). Bronchoalveolar lavage fluid and lung tissue samples were obtained on day 7 after inoculation to assess lung inflammation, airway reactivity, and expression of cytokines and BRP-39. Results:In human subjects, YKL-40 and IL-13 levels in NPA were higher in children with RSV infection than in control subjects. Expression of BRP-39 and Th2 cytokines, IL-13 in particular, was increased following RSV infection in mice. Airway inflammation caused by RSV infection was reduced in BRP-39 KO mice as compared to WT mice. Th2 cytokine levels were not increased in the lungs of RSV-infected BRP-39 KO mice. BRP-39 regulated M2 macrophage activation in RSV-infected mice. Additionally, treatment with anti-CHI3L1 antibody attenuated airway inflammation and Th2 cytokine production in RSV-infected WT mice. Conclusion:These findings suggest that CHI3L1 could contribute to airway inflammation induced by RSV infection. CHI3L1 could be a potential therapeutic candidate for attenuating Th2-associated immunopathology during RSV infection. K E Y W O R D Sbronchiolitis, chitinase 3-like 1 protein, lower respiratory tract infection, respiratory syncytial viruses, type 2 immunity