[The relation between morphologic and functional airway changes in bronchial asthma]. / De relatie tussen morfologische en functionele luchtwegveranderingen in bronchiaalastma.

Asthma is currently defined as a chronic inflammatory disorder of the airways. The exact relationship between this inflammatory process and altered airway behavior in asthma remains to be fully established. More specifically, the question remains as to the exact causal relationship between airway inflammation and bronchial hyperresponsiveness (BHR), which constitutes the predominant lung function abnormality in asthma. The two main determinants of BHR are the hypersensitivity and the hyperreactivity of the airways. Hypersensitivity is reflected in a leftward shift of the dose response curve to the bronchoconstrictor effect of agonists such as histamine. More important from a clinical perspective is the hyperreactivity of the airways, which is characterized by excessive airway narrowing. The airway wall consists of three compartments, namely the inner airway wall area between the airway lumen and the smooth muscle layer, the airway smooth muscle layer and the outer airway wall area between the smooth muscle layer and the lung parenchyma. Mathematical models have calculated that changes within each of these compartments can contribute to airway hyperreactivity. Morphometric analysis of asthmatic airways confirm thickening of each of these three airway wall compartments. The contribution of airway inflammation to the thickening of each of these compartments and their relative functional impact on BHR remains to be further elucidated. Asthmatic airways display signs both of the acute and the chronic phase of an inflammatory reaction. The acute allergic inflammation is characterized by the presence of increased amounts of inflammatory cells including eosinophils, mast cells, macrophages, dendritic cells and T helper 2 (Th2) lymphocytes, and is regulated by a complex network of mutually interacting cytokines. The Th2 lymphocyte plays a crucial role within this network. Based, amongst other observations, on in vivo animal models, the hypothesis can be formulated that through the release of a range of cytokines, Th2 cells affect directly airway responsiveness. It would appear that neither crosslinking of IgE and subsequent mast cell degranulation nor eosinophil influx into the airways play a crucial role in this process. Asthmatic airways also display signs of a chronic inflammatory process, that results in more structural alterations, the so-called airway remodeling. This includes increased deposition of collagen and fibronectin, in addition to airway smooth muscle hypertrophy and hyperplasia. In vivo animal models indicate that these structural alterations have a more profound impact on BHR than the acute inflammation. These models also illustrate that depending on the exact extent and location of structural changes throughout the various airway wall compartments, remodeling can enhance but also protect against excessive airway narrowing, despite the presence of acute inflammation. These results illustrate the necessity to take into account the full extent of histological alterations throughout the airway wall, when evaluating the effect of individual cells and cytokines involved in the acute and chronic inflammatory response in asthma.

Murine models of asthma.

In vivo animal models can offer valuable information on several aspects of asthma pathogenesis and treatment. The mouse is increasingly used in these models, mainly because this species allows for the application in vivo of a broad range of immunological tools, including gene deletion technology. Mice, therefore, seem particularly useful to further elucidate factors influencing the response to inhaled allergens. Examples include: the role of immunoregulatory mechanisms that protect against T-helper cell type 2 cell development; the trafficking of T-cells; and the contribution of the innate immunity. However, as for other animal species, murine models also have limitations. Mice do not spontaneously develop asthma and no model mimics the entire asthma phenotype. Instead, mice should be used to model specific traits of the human disease. The present task force report draws attention to specific aspects of lung structure and function that need to be borne in mind when developing such models and interpreting the results. In particular, efforts should be made to develop models that mimic the lung function changes characteristic of asthma as closely as possible. A large section of this report is therefore devoted to an overview of airway function and its measurement in mice.

[Diffuse interstitial lung disorders in systemic diseases]. / Diffuus interstitieel longlijden bij systeemziekten.

Diffuse parenchymal lung disorders (DPLD) can develop in a variety of systemic disorders. Schematically grouped, these include connective tissue disorders, vasculitis, neoplastic disorders, sarcoidosis and a group of inherited or other rare miscellaneous disorders. This overview focuses on sarcoidosis, systemic sclerosis and Churg Strauss vasculitis. Pulmonary involvement occurs in more than 90% of all patients with sarcodosis. Grading into 4 stages is based on the chest radiograph. Forms characterised by an acute clinical onset or a low grade lung involvement have the highest spontaneous remission rate. The cause of sarcoidosis remains unknown. The diagnosis therefore is descriptive, based on the combination of clinical observations, chest X ray, and the histological documentation of non-caseating epitheloid granulomas in tissue biopsies. Treatment with steroids is only indicated if organ involvement leads to functional impairment. Lung fibrosis is the most important complication of both the "limited" and "diffuse cutaneous form" of systemic sclerosis, involving 90% of all patients. The histological pattern is that of "Usual Interstitial Pneumonia" (UIP) or "Non-specific Interstitial Pneumonia" (NSIP). The pathogenesis of the disorder is thought to consist of an abnormal, excessive regenerative response to an auto-immune mediated lung injury. Churg Strauss vasculitis is characterised by asthma, blood eosinophilia and vasculitis of the small vessels. The affected vessels wall shows signs of fibrinoid necrosis and are infiltrated by eosinophils. pANCA (anti-myeloperoxidase) is considered to play a role in the pathogenesis of the disease. Concern has risen that CysLT1 receptor antagonists might induce production of pANCA. To date, this has not been substantiated.

Dose-related effect of inhaled fluticasone on allergen-induced airway changes in rats.

To examine whether fluticasone propionate (FP) dose-dependently inhibits inflammatory as well as structural changes, Brown Norway rats were sensitised to ovalbumin (OA) on day 0 and 7. From day 14-28, rats were exposed to aerosolised OA (1%) or phosphate buffered saline every 2 days. Thirty minutes before each allergen exposure, animals were pre-treated with aerosolised placebo or FP (0.1, 1 or 10 mg) or prednisolone 3 mg x kg(-1) i.p. At day 29, 0.1 mg FP had no measurable effect, either on inflammatory or structural changes, such as goblet cell hyperplasia and airway wall thickening. The allergen-induced increase in eosinophilic inflammation in bronchoalveolar lavage fluid and in the airway mucosa, as well as increased fibronectin deposition, were inhibited by treatment with FP from a dose of 1 mg onwards. Inhibition of goblet cell hyperplasia and thickening of the airway wall required 10 mg inhaled FP. At this dose, systemic effects were observed. However, for a comparable degree of systemic activity, prednisolone was far less effective at preventing airway changes. The dose of inhaled fluticasone propionate required to inhibit allergen-induced structural alterations was higher than to prevent eosinophil influx, and caused systemic side-effects. However, for a similar systemic activity, prednisolone was ineffective in preventing airway remodelling.

Repeated allergen exposure changes collagen composition in airways of sensitised Brown Norway rats.

Increased or altered collagen deposition in the airway wall is one of the characteristics of airway remodelling in asthma. The mechanisms underlying this increase, and its functional consequences remain to be established further. Representative in vivo animal models might be useful in this respect. In the present study, collagen deposition after prolonged allergen exposure was characterised in the airway wall of Brown Norway rats. Sensitised rats were repeatedly exposed to ovalbumin (OA) or phosphate-buffered saline during 2 and 12 weeks. The deposition of collagen type I, III, IV, V and VI was not altered in animals exposed to OA for 2 weeks. After 12 weeks of OA exposure, more collagen type I was deposited in the inner and outer airway wall and more type V and VI collagen was observed in the outer airway wall. At 12 weeks the number of vessels, identified via type IV collagen staining was not increased, but the total vessel area was. In conclusion, prolonged allergen exposure in sensitised rats is associated with enhanced deposition of type I, V and VI collagens and increased vascularity. This suggests that some aspects of airway remodelling in asthma could be driven by long-term allergen exposure.

Fluticasone inhibits the progression of allergen-induced structural airway changes.

BACKGROUND: Inhaled corticosteroids are widely used as first-line therapy in patients with asthma. The concept of early introduction is more and more accepted. OBJECTIVE: In our rat model of airway remodelling, we investigated whether treatment with inhaled fluticasone propionate can inhibit further progression of established structural airway changes. METHODS: Sensitized Brown Norway rats were exposed to aerosolized ovalbumin (1%) from day 14 to 42. From day 28 to 42, animals were treated with inhaled fluticasone or placebo 30 min before each allergen challenge. One control group was exposed to PBS from day 28 to 42, a second control group throughout the whole experiment. RESULTS: Exposure to ovalbumin during 2 weeks induced structural airway changes, including epithelial cell proliferation, increase in airway wall area and fibronectin deposition. Goblet cell number was increased, although not significantly compared with PBS. Continuing allergen exposure for 2 weeks further enhanced each of these features. In addition, the amount of collagen in the airway wall was enhanced by 4 weeks allergen exposure compared with PBS-exposed animals. These additional increases were inhibited by treatment with fluticasone during the last 2 weeks. CONCLUSION: The progression of established allergen-induced structural airway changes in sensitized rats can be inhibited by treatment with fluticasone.

Interleukin-4 and interleukin-5 gene expression and inflammation in the mucus-secreting glands and subepithelial tissue of smokers with chronic bronchitis. Lack of relationship with CD8(+) cells.

We wished to determine if the inflammatory cells surrounding the airway mucus-secreting glands in chronic bronchitis (CB) were associated with interleukin (IL)-4 and IL-5 mRNA expression and whether the CD8 T cell population expressed these cytokines. Digoxigenin-labeled IL-4 and IL-5 antisense RNA probes were used to detect gene expression in 11 asymptomic smokers (AS), 11 smokers with CB alone with normal lung function, and 10 smokers with chronic bronchitis and coexisting chronic obstructive pulmonary disease (CB+COPD; FEV(1)% of predicted of 43-77% and FEV(1)/ FVC of 51-68%). There were approximately three times as many IL-4 than IL-5 mRNA(+) cells. The highest number of IL-4 mRNA(+) cells were in the submucosal glands of the CB group with normal lung function (216/mm(2)), significantly higher than the values in either the AS (63/mm(2)) or the CB+COPD (87/mm(2)) groups, respectively (p < 0.01). There were similar group differences when the total numbers of inflammatory cells were compared. Accordingly, there was a positive correlation between the number of IL-4 mRNA(+) cells and the total number of inflammatory cells in both the subepithelium and glandular compartments (r = 0.60; p = 0.01 and r = 0.70; p = 0.02, respectively). There were no significant associations between the numbers of CD8(+) and IL-4 or IL-5 mRNA(+) cells. Of 1328 IL-4(+) and 1404 CD8(+) cells counted none was double labeled. Of 727 IL-5(+) and 1569 CD8(+) cells, none was double labeled. In contrast, as a positive control, 34% of tumor necrosis factor (TNF)-alpha(+) cells were also CD8(+) and 15% of CD8(+) cells were TNF-alpha positive. Thus, cells other than the CD8(+) phenotype produce IL-4 and IL-5 in CB. We conclude that there is increased inflammation and IL-4 gene expression in the mucus-secreting glands and the airway mucosa of smokers with bronchitis: both are lower in those with CB and coexisting COPD suggesting that airway inflammation in CB is reduced when airway obstruction develops.

New anti-asthma therapies: suppression of the effect of interleukin (IL)-4 and IL-5.

Asthma is currently defined as a chronic inflammatory disorder of the airways. The central role of allergen-specific Th2 cells in the regulation of this mucosal airway inflammation has been highlighted. Hence, there is large interest in the therapeutic potential of an anti-Th2 cell approach. One of the strategies which has been developed, is to inhibit the effect of interleukin (IL)-4 or IL-5, two main Th2 cell derived cytokines. Interleukin-4 is pivotal in the pathogenesis of allergic disorders through its wide range of effects. An important observation, especially during secondary antigen exposure, is the possible redundancy with IL-13. Both cytokines share common elements in their receptor and intracellular signalling pathway. As a result, compounds can be developed that selectively inhibit the effect of either IL-4 or IL-13, or alternatively, by interfering with the common pathway, inhibit the effect of both cytokines. Eosinophils are generally seen as a particularly harmful element in the allergic inflammation. The importance of IL-5 on eosinophil biology has clearly been established. Conversely, in man, the biological effects of IL-5 are largely limited to eosinophil function. Therefore, IL-5 antagonists offer the unique opportunity of selectively neutralizing the effect of eosinophils. Several strategies have now been developed that successfully inhibit the biological effect of interleukin-4 or interleukin-5. Some of these compounds have proven to be biologically active in man. The challenge now is to establish their therapeutic role in asthma.

The allergen-induced airway hyperresponsiveness in a human-mouse chimera model of asthma is T cell and IL-4 and IL-5 dependent.

The cellular and molecular mechanisms involved in the airway hyperresponsiveness (AHR) of patients with allergic asthma remain unclear. A role for Th2 inflammatory cells was suggested based on murine asthma models. No direct evidence exists on the role of these cells in human asthma. The development of a mouse-human chimera might be useful, allowing the in vivo study of the components of the human immune system relevant to asthma. We investigated the role of allergen-reactive T lymphocytes in a human-mouse SCID model. SCID mice were reconstituted intratracheally with human PBMC from healthy, nonallergic, nonasthmatic donors and exposed to an aerosol of house dust mite allergen after i.p. injection with Dermatophagoides pteronyssinus I Ag and alum. The donor T lymphocytes had a Th1 cytokine phenotype. The reconstituted and allergen-challenged mice developed AHR to carbachol. The mouse airways and lungs were infiltrated with human T lymphocytes. No eosinophils or increases in human IgE were observed. The intrapulmonary human T lymphocytes demonstrated an increase in intracytoplasmic IL-4 and IL-5 and a decrease in IFN-gamma after exposure to allergen adjuvant. Antagonizing human IL-4/IL-13 or IL-5 resulted in a normalization of the airway responsiveness, despite a sustained intracellular Th2 cytokine production. These results provide evidence that the activated human allergen-reactive Th2 cells producing IL-4 or IL-5 are pivotal in the induction of AHR, whereas no critical role for eosinophils or IgE could be demonstrated. They also demonstrate that human allergen-specific Th1 lymphocytes can be driven to a Th2 phenotype.

Fluticasone inhibits but does not reverse allergen-induced structural airway changes.

Ethical and technical reasons limit the possibility of evaluating the effects of inhaled corticosteroids on structural changes in airways of humans with asthma. We therefore evaluated whether fluticasone propionate (FP) modifies airway remodeling, induced by repeated allergen exposure in rats. Sensitized BN rats were exposed to aerosolized ovalbumin (OA) for 2 wk. To assess the effect of FP on the development of or on established airway remodeling, animals were treated with aerosolized FP or placebo during allergen exposure or for 2 wk afterward. Compared with animals exposed to phosphate-buffered saline (PBS), OA-challenged animals developed an increase in total airway wall area, enhanced fibronectin deposition, epithelial cell proliferation, goblet cell hyperplasia, and airway hyperresponsiveness. Concomitant treatment with FP decreased all allergen-induced structural changes without being able to reverse them to normal. Initiating FP treatment after the allergen exposure had no effect on any of the OA-induced structural airway changes. The increase in total airway wall area, enhanced fibronectin deposition, and epithelial cell proliferation persisted. The goblet cell hyperplasia disappeared spontaneously. In conclusion, concomitant treatment with FP partly inhibits structural airway changes as well as hyperresponsiveness induced by OA exposure. Post hoc treatment fails to reverse established airway remodeling.

Counterbalancing of TH2-driven allergic airway inflammation by IL-12 does not require IL-10.

BACKGROUND: Asthma is characterized by allergen-induced airway inflammation orchestrated by TH2 cells. The TH1-promoting cytokine IL-12 is capable of inhibiting the TH2-driven allergen-induced airway changes in mice and is therefore regarded as an interesting strategy for treating asthma. OBJECTIVE: The antiallergic effects of IL-12 are only partially dependent of IFN-gamma. Because IL-12 is a potent inducer of the anti-inflammatory cytokine IL-10, the aim of the present study was to investigate in vivo whether the antiallergic effects of IL-12 are mediated through IL-10. METHODS: C57BL/6J-IL-10 knock-out (IL-10(-/-)) mice were sensitized intraperitoneally to ovalbumin (OVA) and subsequently exposed from day 14 to day 21 to aerosolized OVA (1%). IL-12 was administered intraperitoneally during sensitization, subsequent OVA exposure, or both. RESULTS: IL-12 inhibited the OVA-induced airway eosinophilia, despite the absence of IL-10. Moreover, a shift from a TH2 inflammatory pattern toward a TH1 reaction was observed, with concomitant pronounced mononuclear peribronchial inflammation after IL-12 treatment. Allergen-specific IgE synthesis was completely suppressed only when IL-12 was administered along with the allergen sensitization. Furthermore, treating the animals with IL-12 at the time of the secondary allergen challenge resulted not only in a significant suppression of the airway responsiveness but also in an important IFN-gamma-associated toxicity. CONCLUSIONS: These results indicate that IL-12 is able to inhibit allergen-induced airway changes, even in the absence of IL-10. In addition, our results raise concerns regarding the redirection of TH2 inflammation by TH1-inducing therapies because treatment with IL-12 resulted not only in a disappearance of the TH2 inflammation but also in a TH1-driven inflammatory pulmonary pathology.

Cytokines in asthma.

The airway inflammation underlying asthma is regulated by a network of mutually interacting cytokines. The exact functional role of each individual cytokine in the pathogenesis of the disease remains to be fully established. Type 2 T-helper cells are currently considered to play a crucial role in this process. In vivo animal data suggest a sequential involvement of interleukin (IL)-4 and IL-5 in the induction of allergen-induced airway changes. The potential role of other type 2 T-helper cell-like cytokines in asthma is increasingly being recognized. In particular, IL-4 and -13 display a large degree of redundancy. Whereas IL-4 seems to be crucial in the primary allergen sensitization process, IL-13 might be more important during secondary exposure to aerosolized allergen. Animal models also indicate that T-cell-derived cytokine production, rather than eosinophil influx or immunoglobulin-E synthesis, is causally related to altered airway behaviour. An important aspect when evaluating the functional role of cytokines in a complex disease such as asthma is the interaction with other cytokines in the microenvironment. Increased expression of pro-inflammatory cytokines such as tumour necrosis factor-alpha can further enhance the inflammatory process, and is increasingly linked to disease severity. In addition, decreased expression of immunoregulatory cytokines, including interleukin-12, interleukin-18 or interferon gamma could also strengthen the type 2 T-helper cell-driven inflammatory process.

[The role of inflammation in the modulation of bronchial hyperreactivity. Potential therapeutic applications]. / Rôle de l'inflammation dans la modulation de l'hyperréactivité bronchique. Applications thérapeutiques potentielles.

The exact functional contribution of the various inflammatory cells, mediators, cytokines and growth factors present in asthmatic airways to bronchial hyperresponsiveness remains to be fully established. Gene knock-out in vivo animal models can provide valuable information in this respect. Obviously, the closer the animal models mimic human disease, the more relevant this information will be. This constitutes the major limitation of murine asthma models to date. Key characteristics of asthma include from a morphological point of view, signs of an acute allergic airway inflammation in combination with airway remodeling, and from a functional point of view, hypersensitivity and hyperreactivity of the airways. Neither of these two main characteristics are properly mimicked in currently developed animal models. The degree of airway hyperresponsiveness obtained in these models is generally small, when compared to the degree of hyperresponsiveness observed in asthmatics. This probably relates at least in part, to the differences in airway inflammation, as in most of the murine models, only acute inflammatory changes are induced without chronic structural changes that might affect responsiveness to a large degree. The shortcomings of these models notwithstanding, gene knock-out models of asthma have revealed some interesting observations. The majority of these models has evaluated the exact functional role of TH2 cells, interleukin-4, interleukin-5 and IgE in the pathogenesis of allergic airway inflammation and airway hyperresponsiveness. Overall, it can be argued that neither the IgE/mast cell axis, nor the IL-5/eosinophil axis, are the cause of airway hyperresponsiveness, but that the T-cell in its own right is the main determining factor in establishing the degree of bronchial hyperresponsiveness.