Leonard W. Poulter

Lung immunopathology in cases of sudden asthma death

The histopathology of airway inflammation in rare cases of sudden asphyxic asthma death (SAAD) is unclear. This study examines, for the first time, the relative disposition of lymphocyte and macrophage subsets and eosinophils in proximal and distal tissues of such cases. Multiple resection specimens from five cases of SAAD were studied. Tissue blocks were obtained at necroscopy and immediately frozen in liquid nitrogen within 18 hours of death (death occurring within 1 h of the onset of an unprovoked asphyxic asthma attack). After immunohistological staining, frozen sections underwent semi-quantitative analysis (cell counts per unit area) for T-cells, macrophages and eosinophils using computerized imaging systems. Subsets of T-cells and macrophages were estimated using double immunofluorescence techniques. Variability within samples, between samples and between cases was compared. These cases of fatal asthma showed infiltrates of T-cells, macrophages and eosinophils within peribronchial tissues. Distinct from stable asthma, a CD8+ T-cell dominance was found. A high proportion of eosinophils were activated (EG2+), whereas the relative proportion of antigen-presenting cells (RFD1+) did not seem to be abnormal, although numbers of these cells were high. These features were seen both in proximal and distal tissues. The variability of these parameters within an individual was 9.4-15.2%, however, the variability between individual cases was greater. Sudden asphyxic asthma is associated with inflammatory infiltrates both of proximal and distal lung tissues. In contrast to stable asthma, this infiltrate contains large numbers of CD8+ T-cells, suggesting distinct qualitative as well as quantitative characteristics in the immunopathology of sudden asthma death.

Immunological/physiological relationships in asthma: potential regulation by lung macrophages

There is now a consensus that T-cell-mediated inflammation and eosinophil activation in the bronchial wall contribute to the pathogenesis of asthma. However, the relationship between these immunopathological mechanisms and the observed physiological aberrations remain unclear. Here, Len Poulter and colleagues identify the links between T-cell-mediated inflammation and bronchial hyperresponsiveness, and propose a hypothesis for asthma pathogenesis in which the combination of immunological and physiological abnormalities may result in the promotion of disease. Furthermore, they suggest that an integral factor in the prevention of this process is the regulation of bronchial T-cell reactivity by a population of immunosuppressive macrophages.

Cells and cytokines in chronic bronchial infection.

Bronchiectasis is a chronic pulmonary disease with irreversible dilatation of one or more bronchi.’ It is usually associated with chronic production of purulent infected sputum. Although a classic treatise on the histology of bronchiectasis was written 40 years ago,? it is only recently that the cellular immune response within the bronchial wall has been demonstrated.’ The pathogenesis of bronchiectasis is still not entirely clear. It has been proposed that initially a genetic and/or environmental impairment of mucus clearance’,“.’ allows microorganisms to persist sufficiently long in the bronchial tree to establish themselves and produce e x o t ~ x i n s . ~ ~ ~ ~ ~ Their persistence in a normally sterile site stimulates an inflammatory response that fails to eliminate the bacteria but causes “innocent bystander” lung damage, leading to a “vicious circle” of events. Part of this inflammatory response is a considerable traffic of neutrophils to the lumen of the affected bronchial tree,x neutrophils rarely being found in healthy bronchial tissue or lumen. Because of their potential for damaging these neutrophils may be the most significant component of the inflammatory response. The neutrophil influx is probably initiated by chemoattractant factors, which may be of bacterial’’ or host origin (e.g.. complement components).’?

The immunological component of the cellular inflammatory infiltrate in bronchiectasis.

Immunohistological analysis of bronchial biopsy specimens from nine patients with bronchiectasis and four control subjects was performed with a panel of monoclonal antibodies selected to show lymphocyte and macrophage subsets and signs of cellular activation. The cells taking part in the inflammatory response in the bronchial wall of patients with bronchiectasis were almost exclusively mononuclear cells, most of them T lymphocytes. B lymphocytes were observed in biopsy specimens from only two out of nine patients. CD8+ T cells outnumbered CD4+ cells in all patients in a ratio ranging from 2:1 to 10:1. Most T lymphocytes also strongly expressed CD7 antigen and a proportion of them expressed HLA-DR. Most of the lymphocytic infiltration occurred just beneath the basement membrane of the epithelium, though intraepithelial and submucosal infiltration was also seen. Non-lymphoid mononuclear cells expressing the phenotype of dendritic cells and macrophages were found dispersed throughout the infiltrate, most of them expressing HLA-DR. These observations support the hypothesis that cell mediated immunological reactions contribute to the inflammation associated with bronchiectasis.

Allergen-induced CD30 expression on T cells of atopic asthmatics

Background The importance of TH2‐type T cell cytokines in atopic disease is widely accepted. CD30, a member of the TNF/NGF receptor superfamily, is expressed on a proportion of activated CD45RO+ T cells and has been proposed as a marker for TH2 phenotype. CD30 ligand‐CD30 interaction has been shown to positively influence development of the TH2 phenotype, and serum levels of soluble CD30 (sCD30) have been used as prognostic markers in HIV, SLE, Epstein‐Barr Virus infection and Hodgkins Lymphoma but not as yet in allergic disease.

Release of inflammatory mediators from eosinophils following a hyperosmolar stimulus

Airway dehydration and subsequent hyperosmolarity of periciliary fluid are considered critical events in exercise-induced bronchoconstriction (EIB). It has been shown that an in vitro hyperosmolar stimulation of basophils and mast cells with mannitol can induce the release of histamine and leukotrienes. The aim of this study was to establish if a hyperosmolar challenge could trigger activation of eosinophils to release chemokines and lipid mediators. Peripheral blood eosinophils were isolated from seven asthmatic and six non-asthmatic subjects. Hyperosmolar stimulation of eosinophils with mannitol (0.7 M), resulted in a significant increase in LTC4 levels compared to baseline in both asthmatic (15.2+/-4.6 vs. 70.1+/-9.5; P = 0.0002) and control subjects (14.3+/-4.0 vs. 55.6+/-5.6; P = 0.0001). ECP levels did not increase significantly above baseline following mannitol stimulation in either group. This study shows that eosinophils can be activated by a hyperosmolar stimulus. Therefore it seems reasonable to suggest that eosinophils could contribute to EIB.

T-cell cytokines may control the balance of functionally distinct macrophage populations.

As monocytes differentiate into mature macrophages, subsets emerge that exhibit stimulatory, suppressive or phagocytic potential. These functionally distinct subsets can be discriminated using monoclonal antibodies RFD1 and RFD7. As examples of all these subsets have been repeatedly identified within the macrophage pool in a variety of organs the overall functional capacity of this pool will depend on the relative balance of these subpopulations. This study investigates whether this balance present in mature macrophage populations can be regulated by the local influence of T‐cell‐derived cytokines. The dose‐dependent effect of cytokines interferon‐γ (IFN‐γ), interleukins (IL) IL‐2, IL‐4 and IL‐10 on the phenotype and function of monocyte‐derived macrophages was determined. Subsets of mature cells were quantified by identifying RFD1+ RFD7− stimulatory cells (D1+); RFD1− RFD7+ phagocytes (D7+) and RFD1+ RFD7+ suppressive cells (D1/D7+). IFN‐γ and IL‐4 increased the relative proportions of D1+ stimulatory cells and upregulated HLA‐DR expression. IFN‐γ also increased the capacity of the mature macrophage pool to stimulate T‐cell proliferation. IL‐10 reduced the proportions of D1+ stimulatory cells while promoting the differentiation of D7+ phagocytes and D1/D7+ suppressive cells. IL‐10 induced changes also reduced the functional capacity of the mature populations to stimulate T cells in auto and allogenic mixed lymphocyte reactions (MLR). IL‐2 had no effect on differentiation of monocytes. Thus IL‐4 and IFN‐γ are seen to induce the development of stimulatory macrophages while IL‐10 promotes differentiation of monocytes to mature phagocytes and suppressive macrophages. It is concluded that mature macrophage phenotype is ‘plastic’ and under the control of T‐cell‐derived mediators. Furthermore, within the differentiating monocytes, phenotypic change appears to carry with it functional change, thus retaining the relationship between antigen expression and activity in the mature macrophage populations.

Fluticasone propionate induced alterations to lung function and the immunopathology of asthma over time

BACKGROUND Inhaled corticosteroids are the most widely used treatment for asthma, a disease characterised by both functional and immunopathological abnormalities. This study investigated the relative effects of inhaled corticosteroids on these two features of asthma over time. METHODS A randomised, double blind, placebo controlled, parallel group study with inhaled fluticasone propionate, (FP 2 mg daily) was conducted in 27 patients with asthma. Following baseline analysis, the study tested the effects of short term (two weeks) and longer term (eight weeks) treatment. At each time point (0, 2, and 8 weeks) lung function tests were performed and endobronchial biopsy specimens obtained to determine the distribution and number of lymphocyte, macrophage and eosinophil subsets using immunohistological analysis. Twenty three patients completed the study, 11 on FP and 12 placebo. RESULTS FEV1, ΔFEV1, FEF25–75, and FEV1/FVC all improved after two weeks of FP treatment. This improvement was maintained but not increased after eight weeks. PC20FEV1 showed a trend to increase but was not significantly improved at eight weeks. No significant changes were seen in the placebo group. The numbers of T cells, macrophages, and eosinophils in the bronchial wall were reduced by two weeks of treatment with FP but were unaltered by placebo. The improvement offered by FP continued over the eight week period. Reductions in CD4:CD8 ratio and numbers of activated (EG2+) eosinophils were only significant after eight weeks of treatment. CONCLUSIONS These results reveal that FP influences both functional and immunopathological parameters of asthma. Temporal relationships suggest that these are parallel but not necessarily interrelated effects. While short term treatment is effective in “normalising” the functional abnormalities in asthma, the impact of FP on bronchial inflammation appears to be progressive, taking up to eight weeks and more.

Relative effects of inhaled corticosteroids on immunopathology and physiology in asthma: a controlled study.

BACKGROUND: Although corticosteroids are recognised as the most efficacious treatment for bronchial asthma, their mode of action remains unclear. A placebo controlled trial was undertaken of the effect of inhaled corticosteroids on physiological and immmunopathological parameters in asthmatic patients in whom the correlations between these indices were tested after treatment. METHODS: Sixteen patients (two women) with asthma entered a double blind, placebo controlled, parallel study during which they inhaled either budesonide 800 micrograms twice daily or matching placebo for six weeks. Spirometric parameters and bronchial reactivity to histamine and terbutaline were measured and endobronchial biopsy samples were taken before and after treatment. Patients recorded morning and evening flow rates during the treatment period. The biopsy samples were subjected to immunohistological analysis to determine the disposition of inflammatory cells within the bronchial wall. RESULTS: Treatment with budesonide resulted in a significant improvement in the 25-75% forced expiratory flow (FEF25-75) from a mean of 133 l/min before treatment to 169 l/min after treatment, and in the morning peak expiratory flow rate (PEFR) from a mean of 384 l/min before treatment to 415 l/min after treatment. No changes were seen in the placebo group. Comparison between the changes in the immunopathological indices after six weeks of treatment with placebo or budesonide showed a significant reduction in the numbers of mast cells (0.5/unit area to 0.2/ unit area), activated eosinophils, and the expression of HLA-DR antigens (relative density -1.9 before to 1.02 after treatment) on inflammatory cells in response to treatment with budesonide. Although reductions in the numbers of other inflammatory cells within the bronchial wall were recorded using immunohistological analysis, these changes were not statistically significant. Significant correlations were found between changing immunological indices and lung physiology. CONCLUSIONS: This controlled study shows that inhaled corticosteroids cause improvement in physiological and immunopathological parameters in patients with stable asthma that are not seen with placebo, and that cause and effect relationships may exist between these two measures of disease status.

IFN-γ but not IL-4 T cells of the asthmatic bronchial wall show increased incidence of apoptosis

Background Previous observations have established that IFN‐γ production is depressed in CD4+ T cells from atopic asthmatics compared with non‐asthmatics.