Effects of murine endotoxemia on lymphocyte subsets and clearance of staphylococcal pulmonary infection

In a model of staphylococcal pneumonia initiated during systemic endotoxemia in BALB/c mice, a significant reduction of the number of circulating CD4+ and CD8+ T-lymphocytes, B-lymphocytes, and NK cells, as well as lung-resident total T- and CD4+ T-lymphocytes was demonstrated. Staphylococcus aureus exposure only induced a similar decrease of lymphocyte subsets in the blood. However, the number of lung-resident total T- and CD4+ T-lymphocytes was increased. More viable bacteria were recovered from the lungs of S. aureus-infected mice than from those animals previously treated with lipopolysaccharide (LPS) followed by a staphylococcal challenge. These results indicate that LPS-induced reduction in the number of circulating lymphocyte subsets and lung-resident total T- and CD4+ T-lymphocytes do not increase susceptibility to staphylococcal respiratory infection. Moreover, LPS challenge prior to S. aureus exposure significantly improves clearance of the bacteria in the lung.     

Proliferative expansion and acquisition of effector activity by memory CD4+ T cells in the lungs following pulmonary virus infection

The memory CD4+ T cell response to the respiratory syncytial virus (RSV) attachment (G) protein in the lungs of primed BALB/c mice undergoing challenge pulmonary RSV infection is dominated by effector T cells expressing a single Vbeta-chain, Vbeta14. We have used Vbeta14 expression to examine the kinetics of the activation, accumulation, and acquisition of the effector activity of memory CD4+ T cells responding to pulmonary infection. This analysis revealed that proliferative expansion and effector CD4+ T cell differentiation preferentially occur in the respiratory tract following rapid activation within and egress from the lymph nodes draining the respiratory tract. These findings suggest that, in response to natural infection at a peripheral mucosal site such as the lungs, memory CD4+ T cell expansion and differentiation into activated effector T cells may occur predominantly in the peripheral site of infection rather than exclusively in the lymph nodes draining the site of infection.     

Induction of anti-chlamydial mucosal immunity by transcutaneous immunization is enhanced by topical application of GM-CSF

Transcutaneous immunization (TCI) involves the direct application of antigen plus adjuvant to skin, taking advantage of the large numbers of Langerhans cells and other resident skin dendritic cells, that process antigen then migrate to draining lymph nodes where immune responses are initiated. We have used this form of immunization to protect mice against genital tract and respiratory tract chlamydial infection. Protection was associated with local antibody responses in the vagina, uterus and lung as well as strong Th1 responses in the lymph nodes draining the reproductive tract and lungs respectively. In this study we show that topical application of GM-CSF to skin enhances the numbers and activation status of epidermal dendritic cells. Topical application of GM-CSF also increased the immune responses elicited by TCI. GM-CSF supplementation greatly increased cytokine (IFNgamma and IL-4) gene expression in lymph node and splenic cells compared to cells from animals immunized without GM-CSF. IgG responses in serum, uterine lavage and bronchoalveolar lavage and IgA responses in vaginal lavage were also increased by topical application of GM-CSF. The studies show that TCI induces protection against genital and respiratory tract chlamydial infections and that topical application of cytokines such as GM-CSF can enhance TCI-induced antibody and cell-mediated immunity.     

Patterns of lymphatic drainage to individual thoracic and cervical lymph nodes in the rat

Colloidal carbon was used as tracer material to determine the lymphatic drainage to the cervical and thoracic lymph nodes from various regions of the respiratory tract in the F344 rat. While the lung region may be drained mainly to the posterior mediastinal lymph nodes, the tracheal wall drains primarily to the internal jugular and posterior cervical nodes.     

Effect of lincomycin and staphylococcal vaccine on the course of experimental staphylococcal sepsis

Therapeutic efficacy of lincomycin used alone and in combination with inactivated staphylococcal vaccine and the effect of these agents on synthesis of antibodies and their content in blood serum were investigated. Lincomycin was shown to inhibit septic processes in the host. After its administration the number of the pathogens in the blood and organs markedly decreased. At the same time, lincomycin lowered antibody synthesis in the lymphoid organs and the content of alpha-antitoxins in blood serum. The use of lincomycin in combination with inactivated staphylococcal vaccine promoted an increase in the number of the antibody forming cells in the spleen and lymph nodes and the content of the antibodies to the staphylococcal alpha-toxin in blood serum of the animals with staphylococcal sepsis.     

Effect of an autovaccine on the course of an experimental staphylococcal infection]

The effect of autovaccine on the state of cellular immunity in mice with staphylococcal infection was studied. The maximum decrease of staphylococcal dissemination in internal organs, espeically in the lungs, as well as an increase in the intensity of phagocytosis by peritoneal macrophages were observed after the administration of the vaccine by the method of inhalation. The intranasal administration of the vaccine also proved to be more effective than subcutaneous injection. The cumulation of immune response was more pronounced after the aerosol administration of autovaccine, especially in cases of pathological processes in the respiratory organs.     

Uptake of particulate antigens in a nonmammalian lung: phenotypic and functional characterization of avian respiratory phagocytes using bacterial or viral antigens

Major distinctive features of avian lungs are the absence of draining lymph nodes and alveoli and alveolar macrophages (MPhs). However, a large network of MPhs and dendritic cells (DCs) is present in the mucosa of the larger airways and in the linings of the parabronchi. For the modulation of respiratory tract immune responses, for example, by vaccination, a better understanding of Ag uptake in the chicken respiratory tract is needed. In this study, we provide detailed characterization of APCs in chicken lungs, including their functional in vivo activities as measured by the uptake of fluorescently labeled 1-μm beads that are coated with either LPS or inactivated avian influenza A virus (IAV) mimicking the uptake of bacterial or viral Ag. We identified different subsets of MPhs and DCs in chicken lungs, based on the expression of CD11, activation markers, and DEC205. In vivo uptake of LPS- and IAV-beads resulted in an increased percentage MHC class II(+) (MHC II(+)) cells and in the upregulation of CD40. The uptake of LPS-beads resulted in the upregulation of CD80 and MHC II on the cell surface, suggesting either uptake of LPS- and IAV-beads by different subsets of phagocytic cells or LPS-mediated differential activation. Differences in phagosomal acidification indicated that in chicken lungs the MHC II(+) and CD80(+) bead(+) cell population includes DCs and that a large proportion of beads was taken up by MPhs. LPS-bead(+) cells were present in BALT, suggesting local induction of immune responses. Collectively, we characterized the uptake of Ags by phagocytes in the respiratory tract of chickens.     

Characteristics of the B lymphocytes participating in the reaction of allogeneic stem cell inactivation

In this work the B-cells of mouse lymph nodes are characterized. B-cells produce a helper effect on the capacity of the T-lymphocytes of the lymph nodes for inactivating nonsyngeneic stem cells. The study has revealed that the genetic heterogeneity of the B-lymphocytes does not lead to the abolition of their helper activity. B-lymphocytes of "B-mice" have also been shown to be capable of enhancing the inactivating activity of T-cells.     

Respiratory Dendritic Cell Subsets Differ in Their Capacity to Support the Induction of Virus-Specific Cytotoxic CD8+ T Cell Responses

Dendritic cells located at the body surfaces, e.g. skin, respiratory and gastrointestinal tract, play an essential role in the induction of adaptive immune responses to pathogens and inert antigens present at these surfaces. In the respiratory tract, multiple subsets of dendritic cells (RDC) have been identified in both the normal and inflamed lungs. While the importance of RDC in antigen transport from the inflamed or infected respiratory tract to the lymph nodes draining this site is well recognized, the contribution of individual RDC subsets to this process and the precise role of migrant RDC within the lymph nodes in antigen presentation to T cells is not clear. In this report, we demonstrate that two distinct subsets of migrant RDC - exhibiting the CD103⁺ and CD11b^hi phenotype, respectively - are the primary DC presenting antigen to naïve CD4⁺ and CD8⁺ T lymphocytes in the draining nodes in response to respiratory influenza virus infection. Furthermore, the migrant CD103⁺ RDC subset preferentially drives efficient proliferation and differentiation of naive CD8⁺ T cells responding to infection into effector cells, and only the CD103⁺ RDC subset can present to naïve CD8⁺ T cells non-infectious viral vaccine introduced into the respiratory tract. These results identify CD103⁺ and CD11b^hi RDC as critical regulators of the adaptive immune response to respiratory tract infection and potential targets in the design of mucosal vaccines.     

Antigen-specific regulatory T cells develop via the ICOS-ICOS-ligand pathway and inhibit allergen-induced airway hyperreactivity

Asthma is caused by T-helper cell 2 (Th2)-driven immune responses, but the immunological mechanisms that protect against asthma development are poorly understood. T-cell tolerance, induced by respiratory exposure to allergen, can inhibit the development of airway hyperreactivity (AHR), a cardinal feature of asthma, and we show here that regulatory T (T(R)) cells can mediate this protective effect. Mature pulmonary dendritic cells in the bronchial lymph nodes of mice exposed to respiratory allergen induced the development of T(R) cells, in a process that required T-cell costimulation via the inducible costimulator (ICOS-ICOS-ligand pathway. The T(R) cells produced IL-10, and had potent inhibitory activity; when adoptively transferred into sensitized mice, T(R) cells blocked the development of AHR. Both the development and the inhibitory function of regulatory cells were dependent on the presence of IL-10 and on ICOS-ICOS-ligand interactions. These studies demonstrate that T(R) cells and the ICOS-ICOS-ligand signaling pathway are critically involved in respiratory tolerance and in downregulating pulmonary inflammation in asthma.     

Differences in control by UV radiation of inflammatory airways disease in na?ve and allergen pre-sensitised mice

Exposure of skin to UV radiation (UVR) prior to allergen exposure can inhibit inflammatory airways disease in mice by reducing effector CD4+ T cells in both the trachea and the airway draining lymph nodes. This study analysed the immunomodulatory properties of UVR delivered to na?ve versus allergen pre-sensitised mice. In a model of inflammatory airways disease, BALB/c mice were sensitised by peritoneal injection of the allergen, ovalbumin (OVA) (20 μg/mouse), in the adjuvant, alum (4 mg/mouse), on days 0 and 14. On day 21, the mice were exposed to aerosolised OVA and 24 h later, proliferative responses by the cells in the airway draining lymph nodes were examined. UVR (8 kJ m(-2)) was administered 3 days prior to first OVA sensitisation (day -3), or OVA aerosol challenge (day 18). UVR before sensitisation reduced immune responses associated with expression of allergic airways disease; seven days after first OVA sensitisation, regulation of OVA-induced proliferation in vitro but not in vivo by CD4+CD25+ cells from UV-irradiated mice was detected. UVR administered to pre-sensitised mice regulated allergen responsiveness by cells from the airway draining lymph nodes only with a sensitisation protocol involving allergen and adjuvant at 5% strength of the original dose (1 μg OVA in 0.2 mg alum/mouse). These results suggest that UVR may modulate allergic airways disease by two mechanisms. The first, and more potent, is by reducing effector cells in respiratory tissues and requires UV delivery prior to sensitisation. The second, associated with administration to pre-sensitised mice, is weaker and is detected when the mice are sensitised with lower levels of allergen and adjuvant.     

Kinetics of T- and B-lymphocytes in the lymph nodes of human fetuses

Investigations of the lymph nodes embryogenesis had mainly an anatomo-histological character. At the present time a new approach is necessary: elucidation of main immunological characteristics of lymphoid elements, occupying lymph nodes already at early stages of ontogenesis. The aim of the investigation was to study marker composition of lymphocytes, occupying the lymph nodes of various regional groups, that are in anatomical and functional connection with the thymus, Waldeyer-Pirogov lympho-epithelial pharyngeal ring, appendix and Peyer's patches. The anterior, mediastinal, ileocecal and deep cervical lymph nodes have been studied in 23 human fetuses 17-28-week-old. Immunological and morphological peculiarities of development have been followed in the groups of the lymph nodes mentioned. According to the expression of superficial markers the character of heterogeneity in T- and B-cell systems and their kinetics during embryogenesis has been stated to be characteristic for each regional group. In all lymph nodes the number of T-lymphocytes predominate, their greatest content is noted in the ileocecal lymph nodes. The B-lymphatic system in the lymph nodes is presented poorly with its predominance among immunoglobulin-positive lymphocytes of Ig M(+)-cells.     

Depletion of CD8+ T cells exacerbates CD4+ Th cell-associated inflammatory lesions during murine mycoplasma respiratory disease

Mycoplasma infection is a leading cause of pneumonia worldwide and can lead to other respiratory complications. A component of mycoplasma respiratory diseases is immunopathologic, suggesting that lymphocyte activation is a key event in the progression of these chronic inflammatory diseases. The present study delineates the changes in T cell populations and their activation after mycoplasma infection and determines their association with the pathogenesis of murine Mycoplasma respiratory disease, due to Mycoplasma pulmonis infection. Increases in T cell population numbers in lungs and lower respiratory lymph nodes were associated with the development of mycoplasma respiratory disease. Although both pulmonary Th and CD8(+) T cells increased after mycoplasma infection, there was a preferential expansion of Th cells. Mycoplasma-specific Th2 responses were dominant in lower respiratory lymph nodes, while Th1 responses predominated in spleen. However, both mycoplasma-specific Th1 and Th2 cytokine (IL-4 and IFN-gamma) responses were present in the lungs, with Th1 cell activation as a major component of the pulmonary Th cell response. Although a smaller component of the T cell response, mycoplasma-specific CD8(+) T cells were also a significant component of pulmonary lymphoid responses. In vivo depletion of CD8(+) T cells resulted in dramatically more severe pulmonary disease, while depletion of CD4(+) T cells reduced its severity, but there was no change in mycoplasma numbers in lungs after cell depletion. Thus, mycoplasma-specific Th1 and CD8(+) T cell activation in the lung plays a critical regulatory role in development of immunopathologic reactions in Mycoplasma respiratory disease.     

Scavenger Receptors SR-AI/II and MARCO limit pulmonary dendritic cell migration and allergic airway inflammation

The class A scavenger receptors (SR-A) MARCO and SR-AI/II are expressed on lung macrophages (MPhis) and dendritic cells (DCs) and function in innate defenses against inhaled pathogens and particles. Increased expression of SR-As in the lungs of mice in an OVA-asthma model suggested an additional role in modulating responses to an inhaled allergen. After OVA sensitization and aerosol challenge, SR-AI/II and MARCO-deficient mice exhibited greater eosinophilic airway inflammation and airway hyperresponsiveness compared with wild-type mice. A role for simple SR-A-mediated Ag clearance ("scavenging") by lung MPhis was excluded by the observation of a comparable uptake of fluorescent OVA by wild-type and SR-A-deficient lung MPhis and DCs. In contrast, airway instillation of fluorescent Ag revealed a significantly higher traffic of labeled DCs to thoracic lymph nodes in SR-A-deficient mice than in controls. The increased migration of SR-A-deficient DCs was accompanied by the enhanced proliferation in thoracic lymph nodes of adoptively transferred OVA-specific T cells after airway OVA challenge. The data identify a novel role for SR-As expressed on lung DCs in the down-regulation of specific immune responses to aeroallergens by the reduction of DC migration from the site of Ag uptake to the draining lymph nodes.     

Lymphocyte subpopulations in sensitization and specific immunotherapy in an experiment. Lymphocyte subpopulations in the specific immunotherapy of microbial allergy and subsequent heterologous pollen sensitization

The influence exerted by the specific immunotherapy (SIT) of delayed hypersensitivity (DH) to staphylococci and subsequent sensitization with tarragon pollen on the level of immunocompetent cells in the blood and lymphoid organs of guinea pigs was studied. On the whole, SIT normalized the characteristics of T- and B-lymphocytes, altered as the result of experimentally induced DH: the content of T-cells in the peripheral blood and the lymph nodes increased, while the number of B-cells in the blood and T gamma-suppressors increased. The subsequent heterologous sensitization with pollen abolished the effect of SIT, inducing the general decrease of the level of T gamma-lymphocytes and enhancing the number of T-lymphocytes in the lymph nodes.     

A mouse model for in vivo tracking of the major dust mite allergen Der p 2 after inhalation

Inhaled environmental antigens, i.e. allergens, cause allergic symptoms in millions of patients worldwide. As little is known about the fate of an allergen upon inhalation, we addressed this issue for a major dust mite allergen, Der p 2. First, a model for Der p 2-sensitization was established in C57BL/6 J mice, in which sensitized mice mounted a Der p 2-specific IgE-response with eosinophilic lung inflammation after allergen challenge in the airways. In this model, we applied recombinant Der p 2 carrying a novel C-terminal tetrapeptide Sel-tag enabling labelling with the gamma-emitting radionuclide 75Se at a single selenocysteine residue ([75Se]Der p 2). In vivo tracking of intratracheally administered [75Se]Der p 2 using whole-body autoradiography revealed that [75Se]Der p 2-derived radioactivity persisted in the lungs of sensitized mice as long as 48 h. Radioactivity was also detected in kidneys, liver and in enlarged lung-associated lymph nodes. Interestingly, a larger proportion of radioactivity was found in the lungs of sensitized compared with nonsensitized mice 24 h after intratracheal instillation of [75Se]Der p 2. A radioactive protein corresponding to intact Der p 2 could only be detected in the lungs, whereas [75Se]Der p 2-derived radioactivity was recovered in known selenoproteins both in lung and other organs. Hence, using the recently developed Sel-tag method in a mouse model for Der p 2-sensitization, we could track the fate of an inhaled allergen in vivo. Based upon our findings, we conclude that the inflammatory state of the lung influences the rate of metabolism and clearance of Der p 2. Thus, an allergic response to the inhaled allergen may lead to prolonged retention of Der p 2 in the lung.     

Unusual distribution of Ia-like antigens on canine lymphocytes

The murine monoclonal antibody 7.2, specific for a framework determinant of human Ia antigens, cross-reacts with canine cell membranes recognizing a bimolecular complex (29 000 and 34 000 daltons) similar to that described in man. We investigated the distribution of these Ia-like antigens on mononuclear cells in peripheral blood, thoracic-duct lymph, marrow, alveolar lavage fluids, lymph nodes, and thymuses from normal dogs. By complement-mediated cytotoxicity and indirect immunofluorescence, virtually all lymphocytes expressing surface immunoglobulin (B lymphocytes), monocytes/macrophages, dendritic cells, and many thymus-epithelial cells were la-positive. Furthermore, most non-B-lymphocytes in peripheral blood, thoracic-duct lymph, and lymph nodes expressed Ia antigens. Alveolar (T) lymphocytes and most thymocytes were la-negative. Generally, fluorescence intensity was higher on monocytes/macrophages and B lymphocytes than on non-B-lymphocytes. In mixed leukocyte cultures and concanavalin A-induced blastogenesis assays, treatment of responder cells with antibody 7.2 and complement abolished proliferation. Proliferative responses could not be restored by adding untreated accessory cells, indicating that cytolytic treatment had eliminated responder T-lymphocytes. However, addition of antibody alone to cultures had no significant effect. These studies indicate that most mature canine T-lymphocytes express la-like antigens. Whether this is an intrinsic property of canine cells or possibly related to continuous in vivo stimulation remains to be determined.     

Cytotoxic T-lymphocyte epitope vaccination protects against human metapneumovirus infection and disease in mice

Human metapneumovirus (hMPV) has emerged as an important human respiratory pathogen causing upper and lower respiratory tract infections in young children and older adults. In addition, hMPV infection is associated with asthma exacerbation in young children. Recent epidemiological evidence indicates that hMPV may cocirculate with human respiratory syncytial virus (hRSV) and mediate clinical disease similar to that seen with hRSV. Therefore, a vaccine for hMPV is highly desirable. In the present study, we used predictive bioinformatics, peptide immunization, and functional T-cell assays to define hMPV cytotoxic T-lymphocyte (CTL) epitopes recognized by mouse T cells restricted through several major histocompatibility complex class I alleles, including HLA-A*0201. We demonstrate that peptide immunization with hMPV CTL epitopes reduces viral load and immunopathology in the lungs of hMPV-challenged mice and enhances the expression of Th1-type cytokines (gamma interferon and interleukin-12 [IL-12]) in lungs and regional lymph nodes. In addition, we show that levels of Th2-type cytokines (IL-10 and IL-4) are significantly lower in hMPV CTL epitope-vaccinated mice challenged with hMPV. These results demonstrate for the first time the efficacy of an hMPV CTL epitope vaccine in the control of hMPV infection in a murine model.     

Mouse respiratory tract dendritic cell subsets and the immunological fate of inhaled antigens

It is widely accepted that tissue dendritic cells (DC) function as immune sentinels by alerting T cells to foreign antigen after delivering and presenting it in the draining lymph nodes. Over the last two decades, studies in animal models, particularly rodents, have demonstrated that respiratory tract DC are crucial for the adaptive immune response to inhaled antigen. Indeed, the fate of inhaled antigen is inextricably linked to the function of respiratory tract DC. In this review, we will discuss the characteristics of respiratory tract DC from mice and recent data that may help to explain their role in the fate of inhaled antigen.