Bronchus- and nasal-associated lymphoid tissues

Summary: The bronchus‐associated lymphoid tissue (BALT) and the nasal‐associated lymphoid tissue (NALT) constitute organized lymphoid aggregates that are capable of T‐ and B‐cell responses to inhaled antigens. BALT, located mostly at bifurcations of the bronchus in animals and humans, is present in the fetus and develops rapidly following birth, especially in the presence of antigens. Humoral immune responses elicited by BALT are primarily immunoglobulin A secretion both locally and by BALT‐derived B cells that have trafficked to distant mucosal sites. Similarly located T‐cell responses have been noted. On the basis of these findings, the BALT can be thought of as functionally analogous to mucosal lymphoid aggregates in the intestine and is deemed a member of the common mucosal immunologic system. NALT has been described principally in the rodent nasal passage as two separate lymphoid aggregates. It develops after birth, likely in response to antigen, and B‐ and T‐cell responses parallel those that occur in BALT. It is not known whether NALT cells traffic to distant mucosal sites, although mucosal responses have been detected after nasal immunization. NALT appears from many studies to be a functionally distinct lymphoid aggregate when compared with BALT and Peyer's patches. It may exist, however, in humans as a diffuse collection of isolated lymphoid follicles.     

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.     

Nasal immunity to staphylococcal toxic shock is controlled by the nasopharynx-associated lymphoid tissue

The nasopharynx-associated lymphoid tissue (NALT) of humans and other mammals is associated with immunity against airborne infections, though it is generally considered to be a secondary component of the mucosa-associated lymphoid system. We found that protective immunity to a virulence factor of nasal mucosa-colonizing Staphylococcus aureus, staphylococcal enterotoxin B (SEB), requires a functional NALT. We examined the role of NALT using intranasal (IN) vaccination with a recombinant SEB vaccine (rSEBv) combined with an adjuvant in a mouse model of SEB-induced toxic shock. The rSEBv was rapidly internalized by NALT cells at the mucosal barrier, and transport into NALT was accelerated by inclusion of a Toll-like receptor 4 (TLR4) agonist. Vaccine-induced germinal centers of B cells formed within NALT, accompanied by elevated levels of IgA(+) and IgG(+) cells, and these were further increased by TLR4 activation. The NALT was the site of specific anti-rSEBv IgA and IgG production but was also influenced by intraperitoneal (IP) inoculation and perhaps other isolated lymphoid follicles observed within the nasal cavity. Vaccination by the IN route generated robust levels of anti-rSEBv IgA in saliva, nasal secretions, and blood compared to much lower levels after IP vaccination. IN vaccination also induced secretion of anti-rSEBv IgG in the blood and nasal secretions. Significantly, the efficacy of IN vaccination was dependent on NALT, as surgical removal resulted in greater sensitivity to IN challenge with wild-type SEB. Thus, protective immunity to SEB within the nasal sinuses was elicited by responses originating in NALT.     

Histological studies on the ontogeny of bovine gut-associated lymphoid tissue: appearance of T cells and development of IgG+ and IgA+ cells in lymphoid follicles

The development and distribution of lymphocyte subsets in bovine gut-associated lymphoid tissues (ileal and jejunal Peyer's patches (PP)) were examined. Before birth, the composition of lymphocyte subsets in both PP follicles did not differ except for the dimensions of the interfollicular area and the dome region. Many IgM+ cells were observed in these follicles, but very few CD3+, IgG+, and IgA+ cells could be found. At neonatal period, the IgG+ cells, which did not produce IgG mRNA, were dominant within both PP follicles. From 1 month after birth, many CD3+ cells, IgG mRNA expression, and IgA mRNA expression were detected within the jejunal PP follicles, but very few were in the ileal PP follicles. These data suggest that the characteristics of the jejunal PP follicles metamorphose into secondary lymphoid tissue such as germinal centers at around 1 month after birth, whereas the characteristics of ileal PP follicles were distinct from those of germinal centers.     

Nasal lymphoid tissue, intranasal immunization, and compartmentalization of the common mucosal immune system

Mucosal application of vaccines with an appropriate adjuvant can induce immune responses at both systemic and mucosal sites, and therefore may prevent not only infectious disease, but also colonization at mucosal surfaces. Intranasal is more effective than intragastric immunization at generating earlier and stronger mucosal immune responses. Nasal lymphoid tissue (NALT) and its local draining lymph nodes may retain long-term immune memory. IgA isotype switching, and the differentiation and maturation of IgA antibody-secreting cells (ASC) may occur before these cells migrate out of NALT, whereas IgG ASC responses require passage of the cells through draining lymph nodes of the NALT. Knowledge of whether immune memory cells can recirculate to and reside in the inductive sites other than their origin after encountering antigen will be helpful for understanding the compartmentalization of the common mucosal immune system as well as for determining the best route for delivering a mucosal vaccine against a particular pathogen.     

Reoviruses as probes of the gut mucosal T cell population

Reovirus, serotype 1, causes a transient, asymptomatic infection of the murine intestine when given intraduodenally or orally. However, this infection markedly perturbs both B- and T- cell populations in Peyer's patches (PP) resulting in: 1) a rapid and persistent increase in specific precursors for cytotoxic T cells (pCTL) and a gradient of frequencies highest in PP and lowest in distal lymphoid tissue; 2) a similar increase in memory B cells committed to IgA; 3) the transient appearance of a subset of germinal center B cells identified by MAb, GC-T; 4) the appearance of pCTL among intraepithelial lymphocytes; and 5) the antigen non-specific alteration in Ig isotype potential of B cells previously primed and found in PP. The pCTL appearing upon acute gut mucosal infection with reovirus are Thyl+, Lyt2+, virus-specific, viral serotype non-specific, class I MHC haplotype restricted and occur within the subset of T cells which newly appears also identified by MAb GC-T. Infections of both neonatal and severe-combined immunodeficient mice indicate that the elements of the immune system may operate at many levels to resist, limit, contain, and resolve viral infection.     

Immunology and Immunopathology of the Intestines: Reoviruses as Probes of the Gut Mucosal T Cell Population

Reovirus, serotype 1, causes a transient, asymptomatic infection of the murine intestine when given intraduodenally or orally. However, this infection markedly perturbs both B-and T-cell populations in Peyer's patches (PP) resulting in: 1) a rapid and persistent increase in specific precursors for cytotoxic T cells (pCTL) and a gradient of frequencies highest in PP and lowest in distal lymphoid tissue; 2) a similar increase in memory B cells committed to IgA; 3) the transient appearance of a subset of germinal center B cells identified by MAb, GC-T; A) the appearance of pCTL among intraepithelial lymphocytes; and 5) the antigen nonspecific alteration in Ig isotype potential of B cells previously primed and found in PP. The pCTL appearing upon acute gut mucosal infection with reovirus are Thy1⁺, Lyt2⁺, virus-specific, viral serotype non-specific, class I MHC haplotype restricted and occur within the subset of T cells which newly appears also identified by MAb GC-T. Infections of both neonatal and severe-combined immunodeficient mice indicate that the elements of the immune system may operate at many levels to resist, limit, contain, and resolve viral infection.     

Coincidence of different structures of mucosa-associated lymphoid tissue (MALT) in the respiratory tract of children: no indications for enhanced mucosal immunostimulation in sudden infant death syndrome (SIDS)

Mucosa-associated lymphoid tissue (MALT) is the principal inductive site for mucosal immune responses that are capable of T and B cell responses and antigen-specific responses. In previous independent studies different structures of MALT, e.g. bronchus-, larynx- and nose-associated lymphoid tissue (BALT, LALT, NALT) have been described separately in various frequencies in the human respiratory tract over life spans. Because upper respiratory tract infections are common in infants, dysregulations of mucosal immune responses might be seriously involved in the aetiology of sudden infant death syndrome (SIDS). In the present study the coincidental occurrence of the three different MALT structures in the respiratory tract within the same patients were studied, and cases of SIDS and children who had died from different traumatic and natural causes of death (non-SIDS) were compared. First, the frequency of BALT and LALT in 46 children (35 SIDS, 11 non-SIDS) with or without NALT were examined. A tendency was found of a coincidence of respiratory MALT structures. In 50 additional cases of infant death (30 SIDS, 20 non-SIDS) from the multi-centric German Study on Sudden Infant Death Syndrome (GeSID) where death had occurred in the first year of life, the coincidence was evaluated. A coincidental occurrence of BALT, LALT and NALT or BALT and LALT (each about 30%) was found in both groups, whereby the coincidence in SIDS and the control patients did not differ. Interestingly, the children with coincidental MALT were strikingly older, supporting the hypothesis of respiratory MALT formation via environmental stimulation over time.     

Function of Mucosa-Associated Lymphoid Tissue in Antibody Formation

Abundant evidence supports the notion that human intestinal plasma cells are largely derived from B cells initially activated in gut-associated lymphoid tissue (GALT). Nevertheless, insufficient knowledge exists about the uptake, processing, and presentation of luminal antigens occurring in GALT to accomplish priming and sustained expansion of mucosal B cells. Also, it is unclear how the germinal center reaction so strikingly promotes class switch to IgA and expression of J chain, although the commensal microbiota appears to contribute to both diversification and memory. B-cell migration from GALT to the intestinal lamina propria is guided by rather well-defined adhesion molecules and chemokines/chemokine receptors, but the cues directing homing to secretory effector sites beyond the gut require better definition. In this respect, the role of human Waldeyer's ring (including adenoids and the palatine tonsils) as a regional mucosa-associated lymphoid tissue must be better defined, although the balance of evidence suggests that it functions as nasopharynx-associated lymphoid tissue (NALT) like the characteristic NALT structures in rodents. Altogether, data suggest a remarkable compartmentalization of the mucosal immune system that must be taken into account in the development of effective local vaccines to protect specifically the airways, small and large intestines, and the female genital tract.     

Kinetics of mouse antibody and lymphocyte responses during intranasal vaccination with a lipooligosaccharide-based conjugate vaccine

We investigated the kinetics of humoral immunity and its related cellular immune responses to intranasal (IN) immunization with a detoxified lipooligosaccharide (dLOS)-tetanus toxoid (TT) conjugate against nontypeable Haemophilus influenzae (NTHi) in mice. IN vaccination with dLOS-TT elicited high titers of LOS-specific IgA in nasal washes and IgG in sera during a course of 4 inoculations while high titers of TT-specific IgA and IgG were found in sera. A significant increase of LOS-specific IgA antibody forming cells (AFCs) was observed in nasopharyngeal-associated lymphoid tissue (NALT) and nasal passages. However, TT induced broad responses with higher numbers of IgA and IgG AFCs found in NALT and nasal passages, less but significant IgA AFCs in cervical lymphoid nodes (CLN), spleen, and lungs. Phenotypic analysis revealed a significant rise of total B220+ B-lymphocytes in NALT and CLN, particularly a rise in IgA+/IgM+ cells in the NALT after the immunization. The latter result was complied with a significant rise of IL-4 but not IFN-gamma positive CD4+ T-lymphocytes in NALT. Analysis of IgG antibody subclasses showed that an IgG1 response to both LOS and TT epitopes dominated in serum when compared to IgG2a. These kinetic antibody patterns and cellular responses may provide useful information regarding to effective mucosal vaccines against NTHi infections.     

Use of Peyer''s patch and lymph node fragment cultures to compare local immune responses to Morganella morganii.

Lymphoid tissue fragment cultures were established to analyze the differentiative processes among B cells in Peyer's patches (PP) and peripheral lymph nodes (PLN), especially those in germinal centers. PP cultures from both conventionally reared mice and formerly germ-free mice colonized with Morganella morganii could be maintained for greater than 12 days with continued B-cell division, especially among cells binding high levels of peanut agglutinin, a characteristic of germinal center cells. PLN cultures from conventionally reared mice injected with a heat-killed vaccine of M. morganii could be maintained for the same amount of time. Over this period, PP cultures continued to secrete immunoglobulin A (IgA) as well as smaller amounts of IgM. PP cultures from formerly germ-free mice colonized with M. morganii showed net increases of IgA antiphosphocholine (anti-PC) antibodies with avidities as high as those of the prototypic T15 monoclonal antibody. Similar PLN fragment cultures from conventionally reared mice given footpad injections of M. morganii showed net increases of IgM and IgG anti-PC antibodies in the culture fluid. Thus, although M. morganii stimulated lymphoid tissues in vivo to produce an anti-PC response in vitro when given by either the oral or the parenteral route, the antibody isotypes differed between PP and PLN fragment cultures. Fragment culturing may offer a complementary and simpler way to detect a local secretory IgA response than does either measuring IgA antibody in secretions or detecting IgA antibody in the cytoplasm of plasma cells in the lamina propria of gastrointestinal or respiratory tissue.     

Eosinophils are required to suppress Th2 responses in Peyer's patches during intestinal infection by nematodes

Infections with enteric nematodes result in systemic type 2 helper T (Th2) responses, expansion of immunoglobulin (Ig)G1 antibodies, and eosinophilia. Eosinophils have a supportive role in mucosal Th2 induction during airway hyperreactivity. Whether eosinophils affect the local T-cell and antibody response in the gut-associated lymphoid tissue during enteric infections is unknown. We infected eosinophil-deficient ΔdblGATA-1 mice with the Th2-inducing small intestinal nematode Heligmosomoides polygyrus and found that parasite fecundity was decreased in the absence of eosinophils. A lack of eosinophils resulted in significantly augmented expression of GATA-3 and IL-4 by CD4⁺ T cells during acute infection, a finding strictly limited to Peyer's patches (PP). The increase in IL-4-producing cells in ΔdblGATA-1 mice was particularly evident within the CXCR5⁺PD-1⁺ T-follicular helper cell population and was associated with a switch of germinal centre B cells to IgG1 production and elevated serum IgG1 levels. In contrast, infected wild-type mice had a modest IgG1 response in the PP, whereas successfully maintaining a population of IgA⁺ germinal center B cells. Our results suggest a novel role for eosinophils during intestinal infection whereby they restrict IL-4 responses by follicular T helper cells and IgG1 class switching in the PP to ensure maintenance of local IgA production.     

LOS诱导的特异性抗体分泌细胞的ELISPOT法检测

目的 :动态测定卡他性莫拉氏菌 (Moraxellacatarrhalis,M .cat)脱毒脂寡糖 (dLOS)蛋白质结合疫苗诱导的抗体分泌细胞的应答状态。方法 :以M .catdLOS蛋白质结合疫苗滴鼻免疫BALB/c小鼠。应用酶联免疫斑点试验 (ELISPOT)检测免疫小鼠不同免疫诱导部位和免疫效应部位 ,包括 :鼻相关淋巴组织 (NALT)、脾脏、颈部淋巴结、鼻内容物、肺脏和派伊尔氏结的特异性抗体分泌细胞 ,并同时测定血清、鼻冲洗液、肺泡灌洗液、唾液及粪便提取液中特异性IgA、IgG和IgM的水平。结果 :M .catdLOS蛋白质结合疫苗免疫小鼠的NALT、脾脏、颈部淋巴结、鼻内容物、肺脏和派伊尔氏结中 ,均测出分泌LOS特异抗体的抗体分泌细胞 ,以鼻内容物中IgA分泌细胞的数目最多 ,其次是在NALT和肺脏中 ,这与特异性抗体测定的结果相一致。结论 :M .catdLOS蛋白质结合疫苗经滴鼻免疫 ,能刺激产生LOS特异的黏膜和全身抗体分泌细胞的应答。ELISPOT试验具有快速、灵敏、特异的优点 ,为动态分析单个抗体分泌细胞应答规律提供了新方法。     

Morphological and immunohistochemical studies of the lungs and bronchus-associated lymphoid tissue in a rat model of chronic pulmonary infection with Pseudomonas aeruginosa.

Pseudomonas aeruginosa is one of the most frequently encountered bacterial pathogens in patients with chronic pulmonary infections, including cystic fibrosis and diffuse panbronchiolitis. Bronchus-associated lymphoid tissue (BALT), noted frequently in patients with cystic fibrosis and diffuse panbronchiolitis, is considered to play an important role in the local immunologic defense mechanisms in the respiratory tract. To investigate the role of BALT in chronic pulmonary infections, we developed an animal model for chronic pulmonary infection and studied the morphological and immunohistochemical characteristics of BALT. Experimental pneumonia was produced in rats by intratracheal inoculation of P. aeruginosa enmeshed in agar beads. The histological changes corresponded to those occurring in chronic bronchiolitis. Immunohistochemically, surface immunoglobulin M-positive (sIgM+) cells and sIgA+ cells were recognized in the inflamed bronchial walls from day 4, and sIgG+ cells were recognized from day 14, W3/25+ cells exceeded OX8+ cells in number until day 14. In the BALT, there was a massive accumulation of lymphocytes in the lymphatics and high endothelial venules. The development of germinal centers was accompanied by increased numbers of sIgM+ and sIgA+ cells. W3/25+ cells exceeded OX8+ cells in number in the BALT until day 14. On the other hand, OX8+ cells were predominant in comparison with W3/25+ cells at day 21, and then both sIgM+ and sIgA+ cells and inflammatory changes in the lung decreased at day 28. These findings suggest that BALT regulates the local immune responses against chronic pulmonary infection due to P. aeruginosa.     

A morphologic study of bronchus-associated lymphoid tissue in turkeys

Bronchus-associated lymphoid tissue (BALT) in normal turkeys of ages 1 day and 1, 2, 3, 4, 8, and 18 weeks was examined by light microscopy and by scanning and transmission electron microscopy. Turkey BALT resembled other mucosa-associated lymphoid tissues; it was made up of a population of lymphocytes covered by a specialized epithelium different from typical pseudostratified ciliated columnar bronchial epithelium. There were distinct age-related differences in BALT structure. Bronchus-associated lymphoid nodules were larger and more numerous in older turkeys. In 1-day- to 2-week-old turkeys, the primary cell type of BALT epithelium was nonciliated cuboidal; in 2-week old turkeys it was squamous; and in turkeys older than 4-weeks of age, the epithelium was primarily ciliated columnar. In 1- to 4-week old turkeys, large numbers of intraepithelial lymphocytes disrupted the normal organization of the epithelium. In older turkeys, epithelial and lymphoid cells were in discrete compartments separated by connective tissue. Lymphocytes in 1-day-old turkeys were found in loose aggregates around venules and within the epithelium. In 1-week old turkeys, lymphocytes were organized into compartments of morphologically similar cells. By 3-weeks of age, lymphocytes were present in distinct germinal centers. Epithelial cells of BALT did not have large numbers of apical vesicles and thus were not structurally specialized for antigen uptake by endocytosis. However, the epithelial barrier appeared to be disrupted over lymphoid nodules, suggesting that antigen would be readily available to lymphocytes and phagocytes in BALT. Age-related differences in turkey BALT structure may have functional consequences with respect to the respiratory immune response.     

CD19-deficient mice exhibit poor responsiveness to oral immunization despite evidence of unaltered total IgA levels, germinal centers and IgA-isotype switching in Peyer's patches

CD19 exhibits a critical role as a response regulator in B cells, influencing activation, differentiation and survival. Accordingly, CD19-deficient mice largely lack B-1 cells, and their conventional B-2 cells are poor responders to thymus-dependent antigen. Since both B-1 and B-2 cells may contribute to the total intestinal IgA production, we investigated whether lack of CD19 negatively affected mucosal immunity. We found that CD19(-/-) mice have near normal total IgA levels in serum and gut mucosa and, contrary to systemic lymphoid tissues, Peyer's patches (PP) had germinal centers to which also IgA+ B cells localized. However, the mice demonstrated severely impaired responses to oral immunization with keyhole limpet hemocyanin plus cholera toxin adjuvant. Mucosal responses to oral immunization were significantly more impaired than systemic responses. Despite normal specific IL-4 production, a selective defect in Th2-regulated B cell isotypes was observed, with poor or no mucosal IgA, low serum IgG1 and no IgE, but intact serum IgA and IgG2a production. Ex vivo experiments revealed strongly inhibited CD40-stimulated proliferation and IgA differentiation in CD19-deficient PP B cells. Taken together, an impaired CD40 responsiveness selectively affected Th2, but not Th1, coordinated B cell responses. The CD19(-/-) mice provide compelling evidence for the differential regulation of serum and mucosal IgA immunity.     

Regional differences in the early mucosal immune response induced by primary inoculation of mice with respiratory syncytial virus

Respiratory syncytial virus (RSV) is the most important cause of respiratory tract infection in infants. Little is known about the characteristics of critical immunologic inductive sites within respiratory-associated lymphoid tissues (RALT) upon RSV infection. We examined the kinetics and characteristics of early mucosal RSV-specific immune responses after primary inoculation of mice. We found that the initial production of virus-specific antibodies was restricted to the organized lymphoid tissues of RALT, such as nasal-associated lymphoid tissue (NALT), cervical and bronchial lymph nodes (CLN and BLN). In addition, virus-specific IgM was produced by B cells resident in CLN and BLN, but not NALT, of mice. Finally, we observed regional differences in the pattern of RSV-specific antibodies produced by RALT; B cells within NALT and CLN produced equivalent quantities of virus-specific IgG2a and IgG2b. However, an IgG2a response predominated in BLN. Together these data demonstrate regional differences in the early mucosal immune response to RSV. Further understanding of these differences may assist the development of RSV vaccines.     

Nasal-Associated Lymphoid Tissue is an Inductive Site for Rat Tear IgA Antibody Responses

The role of nasal-associated lymphoid tissue (NALT) as a mucosal inductive site for tear IgA antibody responses was investigated in the rat model. Fluorescent microspheres were shown to access and be taken up by NALT after intranasal or ocular-topical administration, although fewer microspheres were found in the latter case. Tear IgA anti-DNP antibody responses to dinitrophenylated Streptococcus pneumoniae were 6 μg/ml at day 7, 10 μg/ml at day 10, and were still detectable on day 21 (5 μg/ml) following ocular or gastrointestinal immunization. Intranasal immunization induced tear IgA responses which were 1.7-fold higher at day 7 (10 μg/ml), peaked by day 10 (14 μg/ml) and were still 1.6-fold higher (8 μg/ml) at day 21 than responses of ocular or gastrointestinal groups. These findings suggest that intranasal immunization may be more effective than ocular or gastrointestinal administration in eliciting tear IgA antibody responses and, taken together with the microsphere data, indicate that NALT can serve as an inductive site for ocular mucosal IgA responses.     

Structure and function of bronchus-associated lymphoid tissue (BALT)

This article deals with the ontogeny, morphology, and function of bronchus-associated lymphoid tissue (BALT). The surface epithelium of the respiratory tract, including the lymphoepithelium of BALT, is described, with emphasis on infiltrated nonepithelial cells (lymphocytes, macrophages). Special attention is given to antigen handling by the lymphoepithelium and the interaction with BALT and lung macrophages. The structure and function of the diverse types of BALT nonlymphoid cells are discussed. The local immune responses (cellular and humoral reactions), the relationships between local and systemic immune reactions, and the role of BALT in the common mucosal immune system are reviewed. The BALT is compared with Peyer's patches in terms of antigen processing, local immune responses, cell populations, and migration of lymphoid and nonlymphoid cells.