The role of mononuclear phagocytes and dendritic cells in allergic inflammation

Mononuclear-phagocytic system is a diffuse network of cells which includes monoblasts and promonocytes of the bone marrow, blood monocytes, as well as free and fixed tissue macrophage cells. In different tissues and organs macrophages acquire different morphological and functional properties under the influence of the local tissue factors. Interaction of macrophages with other cells and molecules is performed via the large number of different receptors resulting in activation of the macrophage cell, accompanied by a series of morphological and metabolic changes which potentiate all its functions. Activated macrophage cells were found in certain diseases. Macrophages and dendritic cells are associated with all aspects of immunity. Owing to their capacity to undergo phagocytosis they are of the utmost importance for unspecific defense from microorganisms. As accessory cells they also participate in cellular and humoral immunity, being at the same time effector cells owing to their capacity of antigen presentation. Moreover, they also participate in immune response regulation owing to their influence on the function of other cells, including mast cells, basophilic leukocytes and T lymphocytes, in which they may influence differentiation toward Th1 or Th2 and cytokine milieu favorable for allergic reaction. Dendritic cells are the most important antigen-presenting cells and thus, they play a major role in activation of helper T lymphocytes, and mode of antigen presentation is significant for regulation of the nature and intensity of the immune response. Pulmonary macrophage cells have been most thoroughly studied, and the observed changeability of their functional and morphological characteristics is of the utmost importance for studying of the pathogenetic properties and regulation of the chronic inflammatory response in bronchial asthma.     

Comparison of lung dendritic cells and B cells in stimulating naive antigen-specific T cells

Dendritic cells (DCs) are specialized APCs that are important in priming naive T cells and can be manipulated in vitro and in vivo to enhance immunizations against microorganisms and tumors. A limitation in the development of suitable immunotherapeutic vaccines for the lung is incomplete information on the role of DCs and other potential APCs in the lung in priming naive T cells. In the current study, we analyzed the relative contributions of murine lung DCs and B cells to process and present OVA to naive CD4+ OVA323-339-specific (DO11.10) T cells in vitro. We also examined their expression of MHC class II and accessory molecules before and after maturation in culture. Similar to DCs from other sites, freshly isolated lung DCs can process OVA, spontaneously up-regulate MHC class II and accessory molecules during overnight culture, and stimulate naive T cells in an Ag-specific manner. In contrast, freshly isolated lung B cells were unable to both process and present native OVA. Furthermore, under conditions of limited OVA323-339 peptide exposure, B cells had a significantly diminished capacity to stimulate T cells, and this correlated with a decreased density of both MHC class II and important costimulatory molecules as compared with lung DCs.     

Induction of antigen-specific antitumor immunity with adenovirus-transduced dendritic cells

Transduction of dendritic cells (DC) can result in presentation of tumor-associated antigens and induction of immunity against undefined epitopes. The present studies demonstrate adenovirus (Ad)-mediated transduction of the beta-galactosidase gene in mouse DC. Similar transductions have been obtained with the gene encoding the DF3/MUC1 tumor-associated antigen. We show that the Ad-transduced DC are functional in primary allogeneic mixed lymphocyte reactions. Mice immunized with Ad-transduced DC develop cytotoxic T lymphocytes that are specific for the beta-galactosidase or DF3/MUC1 antigens. The results also demonstrate that Ad MUC1-transduced DC induce a specific response which inhibits the growth of DF3/MUC1-positive tumors. These findings support the usefulness of Ad-transduced DC for in vivo immunization against tumor-associated antigens.     

Activation of cutaneous dendritic cells by CpG-containing oligodeoxynucleotides: a role for dendritic cells in the augmentation of Th1 responses by immunostimulatory DNA

Genetic vaccination depends at least in part on the adjuvant properties of plasmids, properties that have been ascribed to unmethylated CpG dinucleotides in bacterial DNA. Because dendritic cells (DC) participate in the T cell priming that occurs during genetic vaccination, we reasoned that CpG-containing DNA might activate DC. Thus, we assessed the effects of CpG oligodeoxynucleotides (CpG ODN) on Langerhans cell (LC)-like murine fetal skin-derived DC (FSDDC) in vitro and on LC in vivo. Treatment with CpG ODN as well as LPS induced FSDDC maturation, manifested by decreased E-cadherin-mediated adhesion, up-regulation of MHC class II and costimulator molecule expression, and acquisition of enhanced accessory cell activity. In contrast to LPS, CpG ODN stimulated FSDDC to produce large amounts of IL-12 but only small amounts of IL-6 and TNF-alpha. Injection of CpG ODN into murine dermis also led to enhanced expression of MHC class II and CD86 Ag by LC in overlying epidermis and intracytoplasmic IL-12 accumulation in a subpopulation of activated LC. We conclude that immunostimulatory CpG ODN stimulate DC in vitro and in vivo. Bacterial DNA-based vaccines may preferentially elicit Th1-predominant immune responses because they activate and mobilize DC and induce them to produce large amounts of IL-12.     

The role of apoptosis in the resolution of T cell-mediated cutaneous inflammation

We have investigated cutaneous purified protein derivative-induced delayed-type hypersensitivity (DTH) responses in healthy volunteers to determine features associated with both the generation and resolution of the reaction. The clinical peak of the response occurred at day 3; however, T cell numbers were maximal on day 7. There was a preferential increase of CD4+ CD45RO+ T cells on day 7, which was largely due to proliferation, since a mean of 19% was in cycle. The proliferation of this subset was associated with the presence of IL-15, which was expressed as early as 12 h, and IL-2, which showed peak expression at 7 days. By day 14, there was a significant decrease in both the mean T cell number/unit area and IL-2 and IL-15 expression in perivascular infiltrates. Maximal CD95 (Fas/Apo-1) ligand and TNF-alpha expression were observed at 7 days and were associated with the presence of 1.83% (range 0.81-2.48%) apoptotic T cells. At 14 days, CD95 ligand and TNF-alpha expression were reduced significantly, and the presence of 2.5% (range 1.5-3.75%) of apoptotic T cells at this time was probably due to cytokine deprivation, associated with decreased Bcl-2 relative to Bax expression. The induction and resolution of the Mantoux reaction may depend on the expression of cytokines, such as IL-2 and IL-15, which regulate both proliferation and apoptosis in T cells. Failure to control either of these phases of the Mantoux reaction may contribute to the chronicity of inflammatory responses in certain cutaneous diseases.     

Cutaneous innervation and the role of neuronal peptides in cutaneous inflammation: a minireview

The patellofemoral joint is a complex articulation based on its dependence on both dynamic and static restraints for stability. Classification of patellofemoral disorders has been frought with confusion. However, progress has been made in the classification and understanding of these disorders by improved understanding of the biomechanics of the joint and by clarification of the terminology to describe patellofemoral pathology. The term chondromalacia patella, although once used as an all-inclusive term for anterior knee pain, is now widely accepted as a term used to describe pathologic lesions of the patellar articular cartilage found at arthroscopy or arthrotomy. An adequate classification system should aid in proper diagnosis and treatment of specific problems. If properly devised, it should also aid in the comparison of results between different treatment centers. In addition, it should be a system that is simple and useful in the clinical setting with minimal use of complicated imaging techniques. From a clinical perspective, patellofemoral problems in the skeletally mature patient fall into three broad categories: 1) patellofemoral instability, i.e., subluxation or dislocation; 2) patellofemoral pain with malalignment but no episodes of instability; and 3) patellofemoral pain without malalignment. The myriad of patellofemoral disorders then fall into subclassifications of these categories. Treatment algorithms can be broadly developed based on the general category, with specific treatments based on the subclassification. In this paper, the authors will present a review of the pertinent literature documenting patellofemoral classification systems and develop concepts of clinical classification of patellofemoral disorders based on the three categories described above.     

Priming and induction of eosinophil trafficking in guinea-pig cutaneous inflammation by tumour necrosis factor alpha

This work addresses the fundamental limits imposed by the MRI process on the accuracy with which vessel diameters and cross-sectional areas can be derived from time-of-flight (TOF) and phase-contrast (PC) MR source images. By means of simulations and in vitro experiments, it is demonstrated that, even in the absence of flow-related artifacts, severe inaccuracies in the determination of diameters or cross-sectional areas may occur solely because of the physical process of the MR image acquisition. Resolution and intraluminal saturation have strong effects on the vessel appearance and thus on the diameter estimation error. It is shown that low resolution leads to diameter overestimation or even underestimation and that intraluminal saturation causes severe underestimation, even for relatively low flip angles. Velocity and velocity encoding do not have a major influence on lumen appearance in PC images. Accurate diameter estimations can be attained only if lumen diameters constitute at least three pixels for both TOF and PC acquisitions, provided that intraluminal saturation is suppressed or avoided. Additionally, since the constitution of TOF and PC images is dissimilar, lumina should be analyzed differently to obtain accurate diameters and cross-sectional areas.     

The role of gammadelta T cells in induction of bacterial antigen-specific protective CD8+ cytotoxic T cells in immune response against the intracellular bacteria Listeria monocytogenes

PURPOSE: To study the relation between abdominal aortic aneurysms and chronical obstructive pulmonary disease (COPD), in particular the suggested common elastin degradation caused by elastase and smoking. METHODS: A cross-sectional population study and a prospective cohort study of small abdominal aortic aneurysms was performed in a community setting. All previous diagnoses recorded in a hospital computer database were received for 4404 men 65 to 73 years of age who had been invited to a population screening for abdominal aortic aneurysm. One hundred forty-one men had AAA (4.2%). They were asked to participate in an interview, a clinical examination, and collection of blood sample. Men with an abdominal aortic aneurysm 3 to 5 cm in diameter were offered annual ultrasound scans to check for expansion. RESULTS: Among patients with COPD 7.7% had abdominal aortic aneurysms (crude odds ratio=2.05). The adjusted odds ratio, however, was only 1.59 after adjustment for coexisting diseases associated with abdominal aortic aneurysm (P=.13). The mean annual expansion was 2.74 mm per year among patients with COPD, 2.72 among patients without COPD, and 4.7 mm among patients who used oral steroids compared with 2.6 among patients who did not use steroids (P < .05). Concentration of serum elastin peptide and plasma elastase-alpha1-antitrypsin complexes correlated negatively with forced expiratory volume in the first second (FEV1) among patients with COPD. However, multivariate regression analysis showed that concentration of serum elastin peptide, therapy with beta-agonists, and FEV1 correlated positively with degree of expansion but that concentration of plasma elastase-alpha1-antitrypsin complexes and serum alpha1-antitrypsin did not influence expansion, suggesting that elastase plays an important role in the pathogenesis of COPD but not of abdominal aortic aneurysm. CONCLUSION: The high prevalence of abdominal aortic aneurysm among patients with COPD is more likely to be caused by medication and coexisting diseases rather than a common pathway of pathogenesis.     

The role of mast cells in the infiltration phenomena in inflammation

On the model of the acute infectious peritonitis in rats it is shown that the previous osmotic depletion of the peritoneal mast cell population leads to the increased functional activity of neutrophils and decreased of monocytes of the inflammatory focus and blood. The results indicate that in the natural inflammatory conditions mast cells directly or indirectly inhibit neutrophils and stimulate monocytes, i.e. they are the modulators of leukocytic reaction.     

Essential role of nuclear factor kappaB in the induction of eosinophilia in allergic airway inflammation

The molecular mechanisms that contribute to an eosinophil-rich airway inflammation in asthma are unclear. A predominantly T helper 2 (Th2)-type cell response has been documented in allergic asthma. Here we show that mice deficient in the p50 subunit of nuclear factor (NF)- kappaB are incapable of mounting eosinophilic airway inflammation compared with wild-type mice. This deficiency was not due to a block in T cell priming or proliferation in the p50(-/-) mice, nor was it due to a defect in the expression of the cell adhesion molecules VCAM-1 and ICAM-1 that are required for the extravasation of eosinophils into the airways. The major defects in the p50(-/-) mice were the lack of production of the Th2 cytokine interleukin 5 and the chemokine eotaxin, which are crucial for proliferation and for differentiation and recruitment, respectively, of eosinophils into the asthmatic airway. Additionally, the p50(-/-) mice were deficient in the production of the chemokines macrophage inflammatory protein (MIP)-1alpha and MIP-1beta that have been implicated in T cell recruitment to sites of inflammation. These results demonstrate a crucial role for NF-kappaB in vivo in the expression of important molecules that have been implicated in the pathogenesis of asthma.     

The role of chemokines and accessory cells in the immunoregulation of cutaneous leishmaniasis

The course of infection with Leishmania parasites is determined by the type of the developing CD4+ T cell immune response. Macrophages and Langerhans cells/dendritic cells play a decisive role in the interaction between the parasites and the host's immune system because they serve as host cells, as accessory cells that present parasite antigen, deliver costimulatory signals and secrete cytokines modulating the T cell activity and as effector cells eliminating the microorganisms. Therefore, we put particular emphasis on characterizing the role of these cells in cutaneous leishmaniasis and the factors regulating their activities. Our results show that (1) expression of the chemokine monocyte chemoattractant protein 1 (MCP-1) is associated with macrophage infiltration into the lesion and stimulation of leishmanicidal activity, (2) Langerhans cells are required for the transport of Leishmania from the infected skin to the draining lymph node and initiation of the specific T cell immune response in the early phase of infection, (3) lymph node dendritic cells containing persistent parasites may be involved in the maintenance of specific immunity, (4) Langerhans cells are able to present L. major LPG to T cells and (5) treatment of mice with antigen-pulsed Langerhans cells induces protective immunity against cutaneous leishmaniasis.     

Efficient presentation of multivalent antigens targeted to various cell surface molecules of dendritic cells and surface Ig of antigen-specific B cells

To study the relation between the form of an Ag and the response to it, we compared presentation in vitro with hen egg lysozyme (HEL)-specific T cells from TCR transgenic mice of free HEL and liposome-encapsulated HEL by different APC. HEL-specific splenic B cells or bone marrow-derived dendritic cells were incubated with free HEL or HEL-containing liposomes targeted by Ab to either surface Ig, the Fc receptor, or MHC class I and II molecules. Ag presentation by HEL-specific B cells was at least 100-fold more efficient for HEL in surface Ig-targeted liposomes than free HEL taken up by the same receptor or HEL in liposomes targeted to class I or II molecules. Ag presentation by dendritic cells from Fc receptor-targeted vesicles was augmented 1,000-10,000-fold compared with free Ag or nontargeted liposomes, but presentation was also efficient when Ag was targeted to class I or II molecules. These results indicate that Ag-specific B cells and dendritic cells can be equally efficient in stimulating IL-2 production by Ag-specific T cells from unimmunized TCR transgenic mice when the Ag is multivalent and taken up by appropriate receptors. In contrast to B cells, which require engagement of surface Ig for optimal presentation, dendritic cells may present Ag by means of several different cell surface molecules.     

The role of T cells in polyethylene particulate induced inflammation

OBJECTIVE: To investigate the role of T lymphocytes in ultra-high molecular weight polyethylene (UHMWPE) induced inflammation in joint arthroplasty. METHOD: We address the role of T cells in wear induced inflammation by injecting the knee joints of both immune competent rats and mice and severe combined immunodeficient (SCID) mice with UHMWPE. Histological and immunohistochemical analysis of the synovial tissues was compared. Interaction between human T cells and UHMWPE particles was examined in vitro using T cell activation assays. RESULTS: Histological and immunohistochemical analysis of the knees of the immune competent animals showed significant UHMWPE induced inflammation. In contrast, the tissue in the SCID mice knee joints showed very little inflammatory response to UHMWPE despite phagocytosis of the particulate. Since the SCID mice have no functional T or B lymphocytes, it is highly likely that the lack of inflammation in knee joints may be due to the absence of mouse T cells, as the infiltration of T cells into the joint tissue may enhance the inflammatory response to UHMWPE particles. T cell activation assays showed that T cells were not directly activated by UHMWPE particles and the nature of the interaction was not revealed from these experiments. CONCLUSIONS: Although T cells are not directly involved in UHMWPE particle induced inflammation, as shown by the T cell activation assays, the histological data from the mice studies clearly show differences in the amplitude of inflammation from animals with and without functional T cells. Our studies suggest that the T cells may enhance the inflammatory response due to a bystander effect. Since the macrophages upon ingestion of UHMWPE particles release several cytokines including tumor necrosis factor-alpha, interleukin 1, and IL-6, it is possible that T cells in the vicinity of these macrophages may become attracted to the knee joint and activated due to cytokine release.     

The role of actin microfilaments in the down-regulation of the degranulation response in RBL-2H3 mast cells

Cross-linking of FcepsilonRI on rat basophilic leukemia (RBL) cells initiates a signaling cascade leading to degranulation of the cells and the release of inflammatory mediators. Inhibitors that disrupt microfilaments, such as latrunculin and cytochalasin D, do not cause any degranulation on their own, but they do enhance FcepsilonRI-mediated degranulation. Dose-response studies show a good correlation between inhibition of actin polymerization and increased degranulation. In RBL cells, latrunculin causes a decrease in basal levels of filamentous actin (F-actin), while cytochalasin D does not. This is particularly evident in the Triton-insoluble pool of F-actin which is highly cross-linked and associated with the plasma membrane. A concentration of 500 nM latrunculin decreases the basal level of Triton-insoluble F-actin by 60-70% and total F-actin levels by 25%. Latrunculin increases both the rate and extent of Ag-induced degranulation while having no effect on pervanadate-induced degranulation. Pervanadate activates the signaling pathways directly and bypasses the cross-linking of the receptor. RBL cells, activated through FcepsilonRI in the presence of latrunculin, show increased phospholipase activity as well as increased tyrosine phosphorylation of Syk and increased tyrosine phosphorylation of the receptor itself by the tyrosine kinase Lyn. This indicates that the very earliest signaling events after receptor cross-linking are enhanced. These results suggest that actin microfilaments may interact, either directly or indirectly, with the receptor itself and that they may regulate the signaling process at the level of receptor phosphorylation. Microfilaments may possibly act by uncoupling Lyn from the cross-linked receptor.     

Expression of complement receptors 1 and 2 on follicular dendritic cells is necessary for the generation of a strong antigen-specific IgG response

Two mechanisms could account for the impaired humoral immune response found in Cr2-/- mice. The absence of complement receptors 1 and 2 (CR1, CR2) on B cells could affect their activation. Alternatively, impaired Ag trapping by follicular dendritic cells (FDC) could affect B cell maturation into Ig-secreting or memory B cells. To compare the roles of CR1 and CR2 on B cells vs FDC in this abnormal response, bone marrow (BM) chimeric mice were generated and immunized with specific T-dependent Ags. The primary and secondary Ab response was measured. Cr2+/+ animals reconstituted with a Cr2-/- BM generated a diminished but detectable humoral immune response compared with controls. When injected with preformed immune complexes (IC), these mice maintained follicular IC localization. Cr2-/- animals reconstituted with a Cr2+/+ BM had an initial rise in the Ab titer, but were unable to maintain it as shown by a pronounced decrease in the IgG titer. This defect persisted during the secondary immune response. Follicular IC trapping was also impaired. Despite the abnormal Ab response, germinal center formation was retained in all of the chimeric animals. These experiments are the first to demonstrate an absolute requirement for CR1 and CR2 expression on FDC in the generation of a normal humoral immune response.     

Studies on the induction of antigen-specific antibody in anti-CD40 cultured human B lymphocytes

Costimulatory signals provided by T cells are required for B cells to produce specific antibody to T-dependent antigen. We have investigated the suitability of using the CD40 culture system for the proliferation and differentiation of Ag-specific human B cells using cytomegalovirus (CMV) or tetanus toxoid (TT) as antigen. We modified the CD40 culture system (CD32-transfected L cells, anti-CD40, and IL-4) by applying a sequential cytokine stimulation and compared total B-cell cultures with antigen-specific B cells preselected by panning. The detection of specific antibody became possible when antigen-selected B cells were cultured for 7 days in the CD40 system to induce clonal expansion, followed by the addition of IL-2 and IL-10 for an additional 7 days to induce plasma-cell differentiation. We conclude that our initial inability to detect specific antibody in the CD40 system is due to overgrowth of nonspecific B-cell clones and that selection of antigen-specific B cells by panning overcomes this problem. Induction of antigen-specific antibody production was found to be optimal when the initial contact with antigen during panning was limited to between 1 to 24 hours.     

In vitro induction of primary, antigen-specific CTL from human peripheral blood mononuclear cells stimulated with synthetic peptides

A protocol for in vitro induction of primary, antigen-specific CTL from human peripheral blood mononuclear cells (PBMCs) was developed. Antigen presenting cells (APCs) consisted of Staphylococcus aureus Cowan-I (SAC-I) activated PBMCs treated with a citrate-phosphate buffer at pH 3 to release endogenous peptides bound to surface MHC. This treatment resulted in transient expression of empty class I molecules which could be subsequently stabilized with peptide and beta 2-microglobulin (beta 2m). SAC-I activated PBMCs from HLA-A2.1 normal donors loaded with HBV core 18-27 peptide following acid treatment were used to stimulate PBMCs depleted of CD4+ T cells, in the presence of recombinant interleukin-7 (rIL-7). After 12 days, cells were restimulated with autologous, peptide-pulsed, adherent cells and tested for CTL activity 7 days later. In 23 independent experiments from 13 different HLA-A2.1 donors, this protocol resulted in induction of primary CTL more than 90% of the time. As indicated by both the frequency and magnitude of the response against peptide-sensitized target cells, SAC-I activated PBMCs treated with acid were the most efficient stimulator APC. Thirteen per cent of the cultures generated were capable of lysing target cells transfected with the HBV core antigen and, in general, these CTL cultures exhibited high avidity for the HBV core peptide. This protocol is generally applicable to different antigens and class I alleles, and thus, may be utilized to screen large numbers of peptides to identify human CTL epitopes.     

IL-2 rescues antigen-specific T cells from radiation or dexamethasone-induced apoptosis. Correlation with induction of Bcl-2

Most studies of apoptosis on T lymphocytes have examined the effects of various stimuli on immature T cells from the thymus. Previous work has indicated that apoptosis of mature memory T cells may be an important pathophysiologic mechanism in diseases such as AIDS, cancer, and autoimmunity. The effect of IL-2 on apoptosis of T cells is not clear. Therefore, we studied the ability of IL-2 to rescue Ag-specific T cells from apoptosis. We found that IL-2, in a dose-dependent manner, prevented T cells from entering apoptosis induced by gamma-irradiation, mitomycin C, or dexamethasone. This effect was specific for IL-2; IL-1 beta, IL-6, or IFN-gamma could not reproduce it. In contrast to Ag-specific T cells, immature T cells and naive mature peripheral T cells could not be rescued by IL-2 from radiation-induced apoptosis. Apoptosis rescue by IL-2 was associated with the induction of bcl-2 mRNA and protein. This induction could not be attributed to the effects of IL-2 on the cell cycle, as T cells that were prevented from cell cycle progression by irradiation showed a similar induction of bcl-2. Rescued cells retained their Ag-specific proliferative capacity and in vivo functions. These findings demonstrate that the apoptotic death of Ag-specific T cell lines, cells which can be regarded as a model for memory T cells, can be prevented with IL-2. This effect may have important therapeutic implications for patients receiving chemotherapy or radiotherapy, and for patients with AIDS who develop immunodeficiency primarily as a result of loss of Ag-specific memory T cells.