Induction of compartmentalized B-cell responses in human tonsils.

The capacity of tonsillar and nasal mucosal lymphoid tissues to serve as induction sites of local and/or distant B-cell responses in humans has been examined. The frequencies of vaccine-specific antibody-secreting cells (ASC) in cell suspensions from palatine tonsils (PT) and adenoids were determined after local (intra-tonsillar [i.t.]) and regional (intranasal [i.n.]) immunizations as well as peroral and parenteral immunizations with cholera and tetanus toxoids. While peroral and parenteral immunizations evoked negligible ASC responses in PT, i.t. vaccination induced a substantial ASC response which consisted of immunoglobulin G (IgG) and IgA ASC. Responses were highly restricted to immunized tonsils. Primary immunization in one PT followed by a second immunization of both PT evoked a larger ASC response in the primed tonsil. The latter ASC response was associated with higher frequencies of ASC precursors in primed tonsils. Furthermore, two i.n. immunizations induced only modest ASC responses in PT, although such immunizations evoked high ASC responses in adenoids. However, both i.t. and i.n. routes of immunization induced specific peripheral blood ASC responses, suggesting that a fraction of B cells activated in tonsils or in nasal mucosa may enter the circulation and disseminate to distant organs. These blood ASC responses preceded increases in both IgA and IgG antibody titers in nasal washes and serum samples. However, vaccine-specific ASC were not detected in duodenal cell suspensions from volunteers who had received i.t. or i.n. immunizations. Collectively, these results indicate that tonsils can serve as expression sites of locally induced antibody responses and support the development of immunological memory. Furthermore, tonsils may serve as powerful inductive sites for immune responses expressed in the upper aerodigestive tract.     

Specific-Antibody-Secreting Cells in the Rectums and Genital Tracts of Nonhuman Primates following Vaccination

To determine optimal strategies to induce specific-antibody-secreting cells (specific ASC) in the rectal and vaginal mucosae, we immunized monkeys with a prototype mucosal immunogen, cholera toxin (CT), given locally or via gastric or parenteral administration. Repeated rectal or vaginal CT immunizations induced strong mucosal and systemic ASC responses. The mucosal responses were, however, confined to the immunization sites and comprised high levels of both specific antitoxin immunoglobulin A (IgA) and IgG. Large numbers of specific IgA and IgG ASC were detected in cell suspensions from dissociated genital and rectal tissues, demonstrating local accumulation of effector B cells at these sites. Intragastric immunization with CT did not per se give rise to cervicovaginal or rectal ASC responses but did prime for a rectal IgA ASC response to local booster immunization. Both rectal and vaginal immunizations also induced circulating blood IgG ASC and IgA ASC. In conclusion, these results show that local administration of antigen to the rectal or vaginal mucosa results in higher ASC responses than systemic or distant mucosal delivery. Furthermore, both the vaginal and the rectal mucosae can serve as inductive sites for systemic ASC responses. These observations should be relevant to the development of vaccines against sexually transmitted diseases such as that caused by human immunodeficiency virus.     

Antibodies and Antibody-Secreting Cells in the Female Genital Tract after Vaginal or Intranasal Immunization with Cholera Toxin B Subunit or Conjugates

We studied the antibody response including antibody-secreting cells (ASC) in the female genital tract of mice after mucosal immunizations with the recombinant B subunit of cholera toxin (rCTB) perorally, intraperitoneally, vaginally, and intranasally (i.n.). The strongest genital antibody responses as measured with a novel perfusion-extraction method were induced after vaginal and i.n. immunizations, and these routes also gave rise to specific immunoglobulin A (IgA) and IgG ASC in the genital mucosa. Specific ASC in the iliac lymph nodes, which drain the female genital tract, were seen only after vaginal immunization. Progesterone treatment increased the ASC response in the genital tissue after all mucosal immunizations but most markedly after vaginal immunization. We also tested rCTB as a carrier for human gamma globulin (HGG) and the effect of adding cholera toxin (CT) as an adjuvant for the induction of systemic and genital antibody responses to HGG after vaginal and i.n. immunizations. Vaginal immunizations with HGG conjugated to rCTB resulted in high levels of genital anti-HGG antibodies whether or not CT was added, while after i.n. immunization the strongest antibody response was seen with the conjugate together with CT. In summary, vaginal and i.n. immunization give rise to a specific mucosal immune response including ASC in the genital tissue, and vaginal immunization also elicits ASC in the iliac lymph nodes. We have also shown that rCTB can act as an efficient carrier for a conjugated antigen for induction of a specific antibody response in the genital tract of mice after vaginal or i.n. immunization.     

Differential expression of chemokine receptors on human IgA+ and IgG+ B cells

Organ-specific lymphocyte homing is dependent on the expression of tissue-specific homing receptors and selected chemokine receptors. During the effector phase of an immune response, IgA and IgG antibody-secreting cells (ASC) are differently distributed in the body. Still, B cell expression of L-selectin and the mucosal homing receptor integrin alpha4beta7 is not related to the isotype produced, but only to the site of antigen encounter. In this study, we examined if differences in chemokine responsiveness between IgA+ and IgG+ B cells could explain their different tissue localization. Circulating CD19+ B cells were isolated and their expression of IgA, IgG, and selected chemokine receptors was determined by flow cytometry. Few Ig+ cells expressed CCR2, CCR3, or CCR9, and there was no difference in the expression of these receptors between IgA+ and IgG+ cells. In contrast, CCR4, CCR5, and CXCR3 was expressed on significantly more IgG+ than IgA+ cells. The function of chemokine receptors on memory B cells and ASC was then tested in the transwell system. IgG+ memory cells migrated to a higher extent than IgA+ cells towards the CXCR3 ligand CXCL11/I-TAC, while there was only a small migration towards the CCR4 ligand CCL17/TARC and the CCR9 ligand CCL25/TECK. ASC migrated poorly to all chemokines tested. In conclusion, this study shows that IgG+ and IgA+ memory B cells have a differential expression of the Th1 associated chemokine receptor CXCR3, as well as of CCR4 and CCR5. In contrast, none of the studied chemokine receptors was preferentially expressed by IgA+ cells.     

Induction of Systemic and Mucosal Immune Responses Following Immunization with Somatostatin-Avidin Complexes Incorporated Into Iscoms

Synthetic, biotinylated somatostatin-14 (Somatotropin Release-Inhibiting Factor; SRIF) was conjugated to avidin, and the resulting complex incorporated into immune-stimulating complexes (ISCOMS). The ISCOMS were used to study the systemic and mucosal immune responses induced by parenteral and gastrointestinal vaccination. Mice were immunized by intraperitoneal (IP) and intragastric (IG) routes and subsequently by either IP or IG secondary immunizations (groups-IP/IP; IP/IG; IG/IG). Antigen specific IgG and IgA antibody secreting cells (ASC) from the spleen, mesenteric lymph nodes (MLN) and Peyer's patches (PP's) were studied by an enzyme-linked immunospot assay (ELISPOT). Specific proliferative responses of spleen cells to avidin and to SRIF were measured.

Immunization IP/IP evoked the highest serum IgG levels to avidin and to SRIF as well as the highest numbers of splenic IgG isotype ASC. The greatest IgA response in MLN and PP's was induced by IP/IG immunization. Only marginal mucosal immunity and no splenic cell specific proliferative responses were found by IG/IG immunization.

These results indicate that ISCOMS are an effective delivery system for protein-peptide antigens. The ISCOMS system described elicited systemic and mucosal antibody immune responses, and primed specific proliferative response when administered IP/IG. This offers another approach for the design and delivery of mucosally administered peptide vaccines.     

Distribution and phenotype of rotavirus-specific B cells induced during the antigen-driven primary response to 2/6 virus-like particles administered by the intrarectal and the intranasal routes

Selection of mucosal sites is an important step in mucosal vaccine development. The intrarectal (IR) route represents an alternative to the oral route of immunization; nevertheless, immune responses induced by this route are not well defined. Here, we studied the early primary B cell response (induction, homing, and phenotype) induced by IR immunization with rotavirus (RV)-2/6 virus-like particles (VLP). Using flow cytometry, we traced RV-specific B cells in different lymphoid tissues and analyzed the expression of alpha4beta7 and CCR9, which are important receptors for homing to the gut, as well as CD5, a marker expressed by B1-a cells, which are a major source of natural antibodies. We observed a massive, specific B cell response in rectal follicles, lumbar, and mesenteric lymph nodes but not in Peyer's patches or cervical lymph nodes. A minority of cells expressed alpha4beta7, suggesting a probable lack of migration to the gut, whereas CCR9 and CD5 were expressed by 30-50% and 30-75% of specific B cells, respectively. Then, we compared the intranasal route of immunization and observed similar B cell frequency and phenotype but in respiratory lymphoid tissues. These results confirm the high compartmentalization of B cell responses within the mucosal system. They show that CCR9 expression, conversely to alpha4beta7, is not restricted to B cells induced in the gut. Finally, an important part of the RV-specific B cell response induced at the mucosal level during the primary response to VLP is most likely a result of B1-a cells.     

Induction of Systemic and Mucosal Immune Responses Following Immunization with Somatostatin-Avidin Complexes Incorporated Into Iscoms

Synthetic, biotinylated somatostatin-14 (Somatotropin Release-Inhibiting Factor; SRIF) was conjugated to avidin, and the resulting complex incorporated into immune-stimulating complexes (ISCOMS). The ISCOMS were used to study the systemic and mucosal immune responses induced by parenteral and gastrointestinal vaccination. Mice were immunized by intraperitoneal (IP) and intragastric (IG) routes and subsequently by either IP or IG secondary immunizations (groups-IP/IP; IP/IG; IG/IG). Antigen specific IgG and IgA antibody secreting cells (ASC) from the spleen, mesenteric lymph nodes (MLN) and Peyer's patches (PP's) were studied by an enzyme-linked immunospot assay (ELISPOT). Specific proliferative responses of spleen cells to avidin and to SRIF were measured.

Immunization IP/IP evoked the highest serum IgG levels to avidin and to SRIF as well as the highest numbers of splenic IgG isotype ASC. The greatest IgA response in MLN and PP's was induced by IP/IG immunization. Only marginal mucosal immunity and no splenic cell specific proliferative responses were found by IG/IG immunization.

These results indicate that ISCOMS are an effective delivery system for protein-peptide antigens. The ISCOMS system described elicited systemic and mucosal antibody immune responses, and primed specific proliferative response when administered IP/IG. This offers another approach for the design and delivery of mucosally administered peptide vaccines.     

Lymphocyte recruitment and protective efficacy against pulmonary mycobacterial infection are independent of the route of prior Mycobacterium bovis BCG immunization

Mycobacterium tuberculosis infects humans through the lung, and immunity to this chronic infection is mediated primarily by CD4(+) T lymphocytes. Recently we have demonstrated that the recruitment of lymphocytes to the lung during primary aerosol M. tuberculosis infection in mice occurs predominantly through the interaction of alpha(4)beta(1) integrin on CD4(+) T cells and vascular cell adhesion molecule-1 on the pulmonary endothelium. To investigate the effect of route of immunization with Mycobacterium bovis BCG on the pattern of T-cell recruitment to the lung, we have analyzed the differences in expression of integrins on activated memory CD4(+) T cells infiltrating the lung following primary BCG immunization by aerosol, intravenous, and subcutaneous routes and after subsequent aerosol challenge with M. tuberculosis. There were marked differences in the patterns of recruitment of activated CD4(+) T cells to the lung following primary immunization by the three routes. Expansion of CD44(hi) CD62L(low) CD4(+) T cells in the lung occurred following aerosol and intravenous BCG immunizations, and the lymphocyte recruitment was proportional to the pulmonary bacterial load. The majority of infiltrating CD4(+) T cells expressed alpha(4)beta(1) integrin. On subsequent exposure to aerosol BCG rapid expansion of gamma interferon-secreting alpha(4)beta(1)(+) CD4(+) T cells occurred to the same extent in all immunized mice, regardless of the route of immunization. Similar expansion of alpha(4)beta(1)(+) CD4(+) memory T cells occurred following M. tuberculosis challenge. The three routes of BCG immunization resulted in the same level of protection against aerosol M. tuberculosis or BCG challenge in both the lungs and spleen. Therefore, recruitment of effector T lymphocytes and protective efficacy against pulmonary mycobacterial infection are independent of the route of prior BCG immunization.     

Cutaneous lymphocyte antigen expression on human effector B cells depends on the site and on the nature of antigen encounter

In contrast to T cells, information on skin-homing B cells expressing the cutaneous lymphocyte antigen (CLA) is sparse. CLA expression on human B cells was investigated among circulating immunoglobulin-secreting cells (ISC) and among antigen-specific antibody-secreting cells (ASC) elicited by parenteral, oral or rectal primary immunization, or by parenteral or oral secondary immunization with Salmonella typhi Ty21a. CLA expression was examined by combining cell sorting with an enzyme-linked immunospot assay. Among all ISC, the proportion of CLA(+) cells was 13-21%. Parenteral immunization induced antigen-specific ASC of which 13% were CLA(+), while oral and rectal immunizations were followed by only 1% of CLA(+) ASC (p<0.001). Oral re-immunization was followed by an up-regulation of CLA (34-48%) regardless of the route of priming. Parenteral re-immunization elicited ASC of which 9-14% were CLA(+). In conclusion, the expression of CLA on human effector B cells depends on the site of antigen encounter: intestinal stimulation elicits cells with no CLA, while parenteral encounter elicits significant numbers of CLA(+) cells. Even though primary antigen encounter in the intestine failed to stimulate CLA expression, up-regulation of CLA was found upon intestinal antigen re-encounter. These findings may be of relevance in the pathogenesis of some cutaneous disorders.     

Topographic distribution of homing receptors on B and T cells in human gut-associated lymphoid tissue: relation of L-selectin and integrin alpha 4 beta 7 to naive and memory phenotypes.

In mice, integrin alpha 4 beta 7 is the main receptor used by lymphocytes that home to the Peyer's patches, although L-selectin contributes to the initial interaction with high endothelial venules. Less is known about the expression and function of these adhesion molecules in humans. The distribution of L-selectin and alpha 4 beta 7 on various B- and T-cell subsets was examined in human Peyer's patches (n = 8) and appendix (n = 4), collectively called gut-associated lymphoid tissue. Multicolor immunophenotyping was performed on cryosections, and dispersed cells were examined by flow cytometry. In cryosections, CD45RA+ T cells around and within interfollicular high endothelial venules, as well as surface (s)IgD+ B lymphocytes in the follicle mantles, often expressed abundant L-selectin but only intermediate levels of alpha 4 beta 7. CD45RO+ T cells and sIgD- B cells expressed higher levels of alpha 4 beta 7 and were often located near putative efferent lymphatics; only a small fraction (< 20%) of such memory cells expressed L-selectin. By flow cytometry, considerably more T than B lymphocytes co-expressed L-selectin and alpha 4 beta 7 (40% versus 25% and 67% versus 39%, respectively). In samples with many L-selectin+ cells (> 30%), more of these lymphocytes co-expressed alpha 4 beta 7 than in samples with few L-selectin+ cells. Because L-selectin and alpha 4 beta 7 were co-expressed on lymphocytes located near high endothelial venules, and because such co-expression was relatively common when many L-selectin+ cells were present, both of these molecules might participate in homing to human gut-associated lymphoid tissue. Such homing is probably most pronounced for T lymphocytes that were found to express L-selectin and alpha 4 beta 7 more often than B lymphocytes. The selective and relatively high expression of alpha 4 beta 7 on memory cells located near efferent lymphatics indicated a different migratory capacity; after exit from gut-associated lymphoid tissue, such stimulated cells might home mainly to mucosal effector sites.     

Cutaneous antibody-secreting cells and B cells in a teleost fish

Antibodies in cutaneous mucus and skin of teleosts play a critical role in the protective immune response against infection. We demonstrate by ELISPOT that antibody-secreting cells (ASC), which include LPS-inducible B cells (plasmablasts) and non-replicating plasma cells, reside in low numbers in the skin of channel catfish. Following immunization against the protozoan parasite Ichthyophthirius multifiliis, which infects skin and gills, the number of ASC in skin increased 20-fold, indicating that the number of ASC in skin is dynamic and increases in response to parasite infection. The number of ASC in skin remained elevated for at least 17 weeks after the last parasite exposure. Cutaneous ASC included I. multifiliis-specific ASC, which undoubtedly serve as the primary source of cutaneous antibodies that confer long-term humoral immunity against reinfection. Our demonstration that skin contains B cells and plasma cells suggests that it is an integral component of the teleost immune system.     

Circulating immunoglobulin-secreting cells are heterogeneous in their expression of maturation markers and homing receptors

Immunoglobulin-secreting cells (ISC) in the peripheral blood are active effectors of the human immune defence system on their way to their site of action. We combined immunomagnetic cell separation and ELISPOT to study the expression of maturation markers and homing receptors (HR) on these cells in healthy volunteers. The results revealed that although highly differentiated, peripheral blood ISC are remarkably heterogeneous both with respect to their expression of maturation markers and HR. Moreover, significant differences were demonstrated between the various isotypes. Fewer IgA-secreting cells expressed both markers of early maturation (HLA-DR, HLA-DQ, CD20, and CD21) and of more mature B cells or plasma cells (CD28, CD38, and α-syndecan) compared with IgG- and IgM-secreting cells. IgA-secreting cells also showed the lowest proportion of cells positive for the peripheral lymph node HR, L-selectin, or the skin HR, cutaneous lymphocyte antigen (CLA). By contrast, the expression of mucosal HR on IgA-secreting cells did not reveal a more pronounced homing attitude to mucosal tissues than IgG- or IgM-secreting cells. We conclude that peripheral blood ISC are a heterogeneous cell population and that IgA-secreting cells seem to differ from the other isotypes both in respect of expression of HR and the various maturational markers studied.     

Antibody-secreting cell responses in the mouse liver.

To elucidate the origins of biliary IgA antibodies, we investigated the isotype and specificity of antibody-secreting cells (ASC) in the liver in comparison with the spleen and intestinal lamina propria of mice immunized by peroral or parenteral routes. The profile of specific IgM, IgG1, IgG2a, and IgA ASC in the liver resembled that of the spleen rather than the lamina propria, regardless of the route of immunization. Peroral immunization increased the proportion of specific IgA ASC in all three organs. However, liver mononuclear cells (MNC) contained a higher proportion of total IgA-secreting cells than spleen cells. After immunization, the number and proportion of B220+ B cells were increased in the liver but not in the spleen. Although the predominant isotype of Ig and specific antibody in bile in response to immunization by either route was IgA, IgM and IgG were clearly detectable. However, specific activities of biliary antibodies relative to total Ig isotype were generally higher than in serum. The predominance of IgA-secreting cells in the liver and the large amount of IgA secreted in the bile resemble the situation at other secretory sites of the mucosal immune system. However, specific antibody-secreting cells appear to accumulate in the liver promptly after immunization, regardless of isotype, and contribute locally produced antibodies to the bile.     

Nasal Lymphoid Tissue (NALT) as a Mucosal Immune Inductive Site

Intranasal (i.n.) immunization is a very effective route for inducing mucosal immunity, but the cellular mechanism responsible for regulating and disseminating these responses is not fully understood. The authors studied the role of nasal lymphoid tissue (NALT) as a mucosal inductive site by using highly purified NALT cells obtained by a new method of mechanical isolation. The NALT cells, like Peyer's patch (PP) cells, were smaller in size and less granular than lymphocytes from salivary glands (SG) and small intestinal lamina propria (LP). The NALT cells isolated from i.n. immunized mice contained antigen-specific antibody-secreting cells (ASC) predominantly of immunoglobulin (Ig)A isotype, similar to those also recovered from salivary glands in a time course study. However, the higher proportion of smaller sized spots formed by NALT cells in ELISPOT assays suggested that these cells were less differentiated precursors of those found in salivary glands. This was supported by the fact that after i.n. immunization, IgA ASC appeared in NALT, as well as in mucosal effector sites SG and LP, but none or very few in another mucosal inductive site, PP. In contrast, after intragastric (i.g.) immunization, IgA ASC were detected in PP, along with the SG and LP, but none or very few in NALT. Furthermore, after i.n. immunization, lymphocytes from NALT but not PP proliferated in response to the specific antigen in culture. These findings imply that NALT served as an inductive site for IgA antibody responses at mucosal effector sites.     

Enhanced mucosal and systemic immune responses to Helicobacter pylori antigens through mucosal priming followed by systemic boosting immunizations

It is estimated that Helicobacter pylori infects the stomachs of over 50% of the world's population and if not treated may cause chronic gastritis, peptic ulcer disease, gastric adenocarcinoma and gastric B-cell lymphoma. The aim of this study was to enhance the mucosal and systemic immune responses against the H. pylori antigens cytotoxin-associated gene A (CagA) and neutrophil-activating protein (NAP), through combinations of mucosal and systemic immunizations in female BALB/c mice. We found that oral or intranasal (i.n.) followed by i.m. immunizations induced significantly higher serum titres against NAP and CagA compared to i.n. alone, oral alone, i.m. alone, i.m. followed by i.n. or i.m. followed by oral immunizations. However, only oral followed by i.m. immunizations induced anti-NAP antibody-secreting cells in the stomach. Moreover, mucosal immunizations alone or in combination with i.m., but not i.m. immunizations alone, induced mucosal immunoglobulin A (IgA) responses in faeces. Any single route or combination of immunization routes with NAP and CagA preferentially induced antigen-specific splenic interleukin-4-secreting cells and far fewer interferon-gamma-secreting cells in the spleen. Moreover, i.n. immunizations alone or in combination with i.m. immunizations induced predominantly serum IgG1 and far less serum IgG2a. Importantly, we found that while both i.n. and i.m. recall immunizations induced similar levels of serum antibody responses, mucosal IgA responses in faeces were only achieved through i.n. recall immunization. Collectively, our data show that mucosal followed by systemic immunization significantly enhanced local and systemic immune responses and that i.n. recall immunization is required to induce both mucosal and systemic memory type responses.     

Phenotypes and distribution of mucosal memory B-cell populations in the SIV/SHIV rhesus macaque model

As vaccine-elicited antibodies have now been associated with HIV protective efficacy, a thorough understanding of mucosal and systemic B-cell development and maturation is needed. We phenotyped mucosal memory B-cells, investigated isotype expression and homing patterns, and defined plasmablasts and plasma cells at three mucosal sites (duodenum, jejunum and rectum) in rhesus macaques, the commonly used animal model for pre-clinical vaccine studies. Unlike humans, macaque mucosal memory B-cells lacked CD27 expression; only two sub-populations were present: naïve (CD21⁺CD27⁻) and tissue-like (CD21⁻CD27⁻) memory. Similar to humans, IgA was the dominant isotype expressed. The homing markers CXCR4, CCR6, CCR9 and α4β7 were differentially expressed between naïve and tissue-like memory B-cells. Mucosal plasmablasts were identified as CD19⁺CD20^+/−HLA-DR⁺Ki-67⁺IRF4⁺CD138^+/− and mucosal plasma cells as CD19⁺CD20⁻HLA-DR⁻Ki-67⁻IRF4⁺CD138⁺. Both populations were CD39^+/−CD27⁻. Plasma cell phenotype was confirmed by spontaneous IgA secretion by ELISpot of positively-selected cells and J-chain expression by real-time PCR. Duodenal, jejunal and rectal samples were similar in B-cell memory phenotype, isotype expression, homing receptors and plasmablast/plasma cell distribution among the three tissues. Thus rectal biopsies adequately monitor B-cell dynamics in the gut mucosa, and provide a critical view of mucosal B-cell events associated with development of vaccine-elicited protective immune responses and SIV/SHIV pathogenesis and disease control.     

Human IgA‐secreting cells induced by intestinal,but not systemic,immunization respond to CCL25 (TECK) and CCL28 (MEC)

Organ‐specific homing of lymphoid cells depends on the expression of tissue‐specific adhesion molecules and production of specific chemokines. CCL25 (TECK) and CCL28 (MEC) have been reported to direct circulating memory/effector B cells to mucosal tissues. Here, we examined if differential responsiveness to mucosal and systemic chemokines could explain the differential migration pattern of circulating human antibody‐secreting cells (ASC), induced by mucosal and systemic immunization. There was a robust migration of specific IgA‐ and IgM‐ASC induced by Salmonella vaccination toward the mucosal chemokines CCL25 and CCL28. In contrast, tetanus‐specific ASC migrated to the systemic chemokine CXCL12 (SDF‐1α) and showed no response to CCL25 or CCL28, not even tetanus‐specific IgA‐ASC. Cell sorting experiments demonstrated that Salmonella‐specific ASC co‐expressed CCR9 and CCR10. Our results show that induction site, rather than isotype commitment, determines the chemokine responsiveness and migration pattern of human effector B cells.     

Specific and Cross‐reactive Plasmablast Response in Humans after Primary and Secondary Immunization with Vi Capsular Polysaccharide Typhoid Vaccine

Secondary immunization with polysaccharide vaccines may imply a risk of hyporesponsiveness. Despite the wide use of typhoid Vi capsular polysaccharide vaccine, its potential tendency to hyporesponsiveness has been inadequately addressed. While previous studies have explored serum antibody responses, we applied a more sensitive approach, a single‐cell assay for circulating plasmablasts, to compare primary and secondary responses. Twelve subjects received primary and booster doses of the Vi vaccine (Typherix^®) at 30‐ to 37‐month intervals. Plasmablasts specific to the Vi or typhoidal O antigens or cross‐reactive with paratyphoid and non‐typhoidal Salmonella strains were identified as antibody‐secreting cells (ASC) with ELISPOT. Before vaccinations, none had plasmablasts specific to the antigens tested. Twelve of 12 subjects showed a Vi‐specific response after primary, but only eight of 12 after booster vaccination. All responded to typhoidal O‐9,12 antigen after both immunizations. The geometric mean of plasmablasts specific to the Vi antigen was 59 (95% CI 24–119) and 1 (0‐54) IgA + IgG + IgM‐ASC/10⁶ peripheral blood mononuclear cell (PBMC) after primary and booster immunizations, respectively, and 20 (9–49) and 56 (29–103) to the O‐9,12 antigen. We detected 1 (0–28) and 17 (6–36) ASC/10⁶ PBMC cross‐reactive with Salmonella Paratyphi A; 3 (0–30) and 22 (8–48) with S. Paratyphi B; 3 (0–29) and 18 (7–47) with S. Paratyphi C; 19 (10‐34) and 51 (26‐94) with Salmonella Enteritidis; and 1 (0–35) and 23 (9–52) with Salmonella Typhimurium, respectively. One‐third of the vaccinees, although responding to the O‐9,12 antigen, failed to respond to the Vi antigen after booster immunization, suggesting hyporesponsiveness in part of the vaccinees. The findings warrant further investigation.     

Mucosal vaccines: a paradigm shift in the development of mucosal adjuvants and delivery vehicles

Mucosal immune responses are the first‐line defensive mechanisms against a variety of infections. Therefore, immunizations of mucosal surfaces from which majority of infectious agents make their entry, helps to protect the body against infections. Hence, vaccinization of mucosal surfaces by using mucosal vaccines provides the basis for generating protective immunity both in the mucosal and systemic immune compartments. Mucosal vaccines offer several advantages over parenteral immunization. For example, (i) ease of administration; (ii) non‐invasiveness; (iii) high‐patient compliance; and (iv) suitability for mass vaccination. Despite these benefits, to date, only very few mucosal vaccines have been developed using whole microorganisms and approved for use in humans. This is due to various challenges associated with the development of an effective mucosal vaccine that can work against a variety of infections, and various problems concerned with the safe delivery of developed vaccine. For instance, protein antigen alone is not just sufficient enough for the optimal delivery of antigen(s) mucosally. Hence, efforts have been made to develop better prophylactic and therapeutic vaccines for improved mucosal Th1 and Th2 immune responses using an efficient and safe immunostimulatory molecule and novel delivery carriers. Therefore, in this review, we have made an attempt to cover the recent advancements in the development of adjuvants and delivery carriers for safe and effective mucosal vaccine production.     

Role of B7 costimulatory molecules in mediating systemic and mucosal antibody responses to attenuated Salmonella enterica serovar Typhimurium and its cloned antigen

The purpose of the present study was to evaluate the ability of an attenuated Salmonella enterica serovar Typhimurium vaccine strain to up-regulate B7-1 and B7-2 on antigen-presenting cells and to examine the functional roles these costimulatory molecules play in mediating immune responses to Salmonella and to an expressed cloned antigen, the saliva-binding region (SBR) of antigen I/II. In vitro stimulation of B cells (B220+), macrophages (CD11b+), and dendritic cells (CD11c+) with S. enterica serovar Typhimurium induced an up-regulation of B7-2 and, especially, B7-1 expression. The in vivo functional roles of B7-1, B7-2, and B7-1/2 were evaluated in BALB/c wild-type and B7-1, B7-2, and B7-1/2 knockout (KO) mice following intranasal immunization with the Salmonella expressing the cloned SBR. Differential requirements for B7-1 and B7-2 were observed upon primary and secondary immunizations. Compared to wild-type controls, B7-1 and B7-2 KO mice had reduced mucosal and systemic anti-Salmonella antibody responses after a single immunization, while only B7-1 KO mice exhibited suppressed anti-Salmonella antibody responses following the second immunization. Mucosal and systemic antibody responses to SBR were reduced following the primary immunization, whereas a compensatory role for either B7-1 or B7-2 was observed after the second immunization. B7-1/2 double KO mice failed to induce detectable levels of mucosal or systemic immunoglobulin A (IgA) or IgG antibody responses to either Salmonella or SBR. These findings demonstrate that B7-1 and B7-2 can play distinct as well as redundant roles for mediating mucosal and systemic antibody responses, which are likely dependent upon the nature of the antigen.