Treatment of airway inflammation in cystic fibrosis

Airway inflammation is now recognized as a major factor in the pathogenesis of cystic fibrosis (CF) lung disease. Therapies aimed at decreasing the inflammatory response represent a new strategy for treatment, and attention has focused primarily on the therapeutic potential of corticosteroids and nonsteroidal anti-inflammatory drugs (NSAIDs). Alternate-day prednisone (1 mg/kg) may be beneficial; however, unacceptable adverse effects limit long-term use. Inhaled corticosteroids are under investigation as a safer alternative. High-dose ibuprofen (approximately 20-30 mg/kg twice daily) has been shown to decrease the progression of CF lung disease, particularly in children with mild lung disease, and it is without significant toxicity. Other NSAIDs (piroxicam) are under consideration, as well as pentoxifylline and fish oil. The rationale for all of these agents lies in their potential to decrease neutrophil influx into the lung. Because of the large burden and deleterious effects of uninhibited neutrophil elastase and oxidants in the CF airway, antiproteases and antioxidants are also being studied. To optimize anti-inflammatory therapy, it is necessary to understand the mechanism of action of these agents in the CF lung, to determine which of these agents would provide the most benefit to patients with CF, and to determine which therapies should be initiated at what age or stage of lung disease. It is hoped that adding anti-inflammatory therapy to an already comprehensive treatment program will decrease morbidity and improve the quality of life for patients with CF.     

Epithelia cells as regulators of airway inflammation

The past decade has seen major advances in our understanding of the metabolic and biochemical functions of the epithelial cell. As a result, the traditional view of the epithelium as a relatively passive physical barrier to the internal environment has been superseded by the concept that the epithelial plays a key role in regulating airway inflammation. The epithelial cell experts this modulatory effect in several ways. Epithelial cells can function maintain mucosal integrity and to modulate local immune responses. They can also limit inflammatory processes by degrading, or inhibiting, proinflammatory mediators and proteins. However, the epithelium also responds to a range of stimuli by producing biologically active mediators that can influence airway inflammation. These include, but are not limited to a broad range of cytokines and chemokines that can exert profound effects on inflammatory cells, as well as lipid and peptide mediators. This review will highlight some of these aspects of the role of the epithelium in regulating airway inflammation.     

The role of rhinovirus in allergic airway inflammation

Epidemiological data demonstrate that viral infections are the most important trigger for acute asthma symptoms in children, and this association persists in many adults with asthma. Studies on volunteers experimentally infected with rhinoviruses (RV) suggest that atopy alone does not predispose to unusually severe symptoms. In contrast, experimental models combining viral infection and allergen exposure have identified potential links between virus-induced and allergen-induced inflammation. While in vitro studies suggest that cytokines may be an important part of this association, their role must be verified by sampling lower airway fluids and tissues in vivo after experimental and/or natural rhinovirus infections. Although it has long been recognized that the common cold is a potent trigger for symptoms of asthma, the mechanisms underlying the association between upper respiratory infection and increased lower airway obstruction remain obscure. The use of experimental infection of volunteers with or without respiratory allergies has enabled direct comparisons of common cold symptoms in these two groups. Furthermore, techniques such as bronchoalveolar lavage and segmental antigen challenge have been used to directly sample lower airway fluids and tissues during acute viral infection.     

Regulation of airway neurogenic inflammation by neutral endopeptidase

Airway neurogenic inflammation is caused by tachykinins released from peripheral nerve endings of sensory neurons within the airways, and is characterized by plasma protein extravasation, airway smooth muscle contraction and increased secretion of mucus. Tachykinins are degraded and inactivated by neutral endopeptidase (NEP), a membrane-bound metallopeptidase, which is located mainly at the surface of airway epithelial cells, but is also present in airway smooth muscle cells, submucosal gland cells and fibroblasts. The key role of NEP in limiting and regulating the neurogenic inflammation provoked by different stimuli has been demonstrated in a large series of studies published in recent years. It has also been shown that a variety of factors, which are relevant for airway diseases, including viral infections, allergen exposure, inhalation of cigarette smoke and other respiratory irritants, is able to reduce NEP activity, thus enhancing the effects of tachykinins within the airways. On the basis of these observations, the reduction of neutral endopeptidase activity may be regarded as a factor that switches neurogenic airway responses from their physiological and protective functions to a detrimental role that increases and perpetuates airway inflammation. However, further studies are needed to assess the role of neutral endopeptidase down regulation in the pathogenesis of asthma and other inflammatory airway diseases.     

Methods for evaluating biological response modifiers

Biological response modifiers (BRM) are a new approach of cancer treatment. The guidelines for their evaluation are comparable to those used for evaluating conventional anticancer treatments. However, it is more difficult to assess their efficacy and toxicity because of their own properties and of the characteristics of underlying diseases. The objectives of studying BRM efficacy are the following : 1- to confirm the pharmacodynamic effect observed in experimental models; 2- to assess the benefit for the patient. Phase II studies can occasionally be performed in healthy volunteers. Efficacy criteria are objective response rate, progression free survival, overall survival or, for hematopoietic growth factors, more specific criteria. In randomized phase III studies patients in the control arm can be observed without treatment, or receive a placebo or a standard treatment. As regards toxicity, side effects specifically related to the use of BRM are the synthesis of cytokines, the stimulation of immune system, the stimulation of tumor growth, the synthesis of anti-mouse of anti-BRM antibodies.     

Time-course of antigen-induced airway inflammation in the guinea-pig and its relationship to airway hyperresponsiveness

The causative relationship between airway inflammation and hyperreactivity is unclear, since inflammatory changes have been examined at one or, at most, a few time-points after antigen challenge in both human asthma and animal models. We have made a detailed investigation of inflammatory and functional changes in the airways up to 8 days after antigen challenge in guinea-pigs. In particular, we examined the hypothesis that eosinophil-derived mediators contribute to tissue damage and the development of airway hyperresponsiveness. Following antigen challenge, the influx of inflammatory cells and mediator release in airway tissue and bronchoalveolar lavage fluid were correlated temporally with histopathological changes in airway tissue and airway responsiveness. Eosinophil influx was demonstrable at 4 h. Eosinophilia peaked after 24 h and persisted for at least 8 days. Parallel increases in the concentrations of major basic protein and eosinophil cationic protein in bronchoalveolar lavage fluid indicated that the eosinophils were activated. Eosinophilia was accompanied by subepithelial oedema and epithelial damage co-localized with major basic protein immunoreactivity. A transient neutrophilia (< 48 h duration) and an increase in neutrophil elastase in bronchoalveolar lavage fluid peaked at 14 h. The proportion of airway macrophages with an activated morphology increased at 8 h and remained markedly elevated until 72 h. Airways were hyperresponsive to histamine at 4 h and for at least 8 days. The antigen-induced airway inflammation resemble in time-course and histopathology that seen in antigen-challenged asthmatics, and indicate that the eosinophil and its cytotoxic proteins may be major mediators of airway mucosal damage and airway hyperresponsiveness.     

Therapies aimed at airway inflammation in cystic fibrosis

Therapies aimed at decreasing the inflammatory response present a new strategy for treating cystic fibrosis (CF) lung disease. Alternate day prednisone may be beneficial, however, unacceptable adverse effects limit long-term use. Inhaled corticosteroids are under investigation as a safer alternative. High-dose ibuprofen twice daily has been shown to decrease the progression of CF lung disease and is without significant toxicity. Other NSAIDs and pentoxifylline and fish oil are under consideration. Antiproteases and antioxidants are also being studied. The rationale for all of these agents lies in their potential to decrease neutrophil influx into the lung, and counteract injurious products of neutrophils. Adding anti-inflammatory therapy to an already comprehensive treatment program will hopefully decrease morbidity and improve the quality of life for patients with CF.     

Requirement for gammadelta T cells in allergic airway inflammation

The factors that contribute to allergic asthma are unclear but the resulting condition is considered a consequence of a type-2 T helper (TH2) cell response. In a model of pulmonary allergic inflammation, mice that lacked gammadelta T cells had decreases in specific immunoglobulin E (IgE) and IgG1 and pulmonary interleukin-5 (IL-5) release as well as in eosinophil and T cell infiltration compared with wild-type mice. These responses were restored by administration of IL-4 to gammadelta T cell-deficient mice during the primary immunization. Thus, gammadelta T cells are essential for inducing IL-4-dependent IgE and IgG1 responses and for TH2-mediated airway inflammation to peptidic antigens.     

Blockade of airway inflammation and hyperresponsiveness by HIV-TAT-dominant negative Ras

We have reported previously that HIV-TAT-dominant negative (dn) Ras inhibits eosinophil adhesion to ICAM-1 after activation by IL-5 and eotaxin. In this study, we evaluated the role of Ras in Ag-induced airway inflammation and hyperresponsiveness by i.p. administration into mice of dnRas, which was fused to an HIV-TAT protein transduction domain (TAT-dnRas). Uptake of TAT-dnRas (t(1/2) = 12 h) was demonstrated in leukocytes after i.p. administration. OVA-sensitization significantly increased eosinophil and lymphocyte numbers in bronchoalveolar lavage fluid 24 h after final challenge. Treatment of animals with 3-10 mg/kg TAT-dnRas blocked the migration of eosinophils from 464 +/- 91 x 10(3)/ml to 288 +/- 79 x 10(3)/ml with 3 mg/kg of TAT-dnRas (p < 0.05), and further decreased to 116 +/- 63 x 10(3)/ml after 10 mg/kg TAT-dnRas (p < 0.01). Histological examination demonstrated that inflammatory cell infiltration (largely eosinophils and mononuclear cells) and mucin production around the airways caused by OVA were blocked by TAT-dnRas. OVA challenge also caused airway hyperresponsiveness to methacholine, which was dose dependently blocked by treatment with TAT-dnRas. TAT-dnRas also blocked Ag-induced IL-4 and IL-5, but not IFN-gamma, production in lung tissue. Intranasal administration of IL-5 caused eosinophil migration into the airway lumen, which was attenuated by pretreatment with TAT-dnRas. By contrast, TAT-green fluorescent protein or dnRas lacking the TAT protein transduction domain did not block airway inflammation, cytokine production, or airway hyperresponsiveness. We conclude that Ras mediates Th2 cytokine production, airway inflammation, and airway hyperresponsiveness in immune-sensitized mice.     

Thromboxane causes airway hyperresponsiveness after cigarette smoke-induced neurogenic inflammation

We investigated the role of neurogenic inflammation and the subsequent mechanisms in cigarette smoke-induced airway hyperresponsiveness in guinea pigs. Exposure to cigarette smoke was carried out at tidal volume for 3 min. Airway responsiveness to histamine was determined before and after smoke exposure followed by bronchoalveolar lavage (BAL). Plasma extravasation was evaluated by measuring the extravasation of Evans blue dye in the airway. Cigarette smoke produced significant airway hyperresponsiveness and plasma extravasation, with an influx of neutrophils in BAL fluid. FK-224 (10 mg/kg i.v.), a tachykinin antagonist at NK1 and NK2 receptors, significantly inhibited these changes. The thromboxane (Tx) B2 concentration was increased in BAL fluid after smoke exposure and was significantly inhibited by FK-224. OKY-046 (10 mg/kg i.v.), a Tx synthase inhibitor, significantly inhibited airway hyperresponsiveness but had no effect on neutrophil influx or plasma extravasation. The results suggest that neurogenic inflammation and the subsequent generation of Tx in the airway are important in the development of the airway hyperresponsiveness induced by cigarette smoke.     

Potential masking effects of salmeterol on airway inflammation in asthma

We hypothesized that regular use of long-acting beta-agonists could delay recognition of ("mask") increasing airway inflammation. We studied steroid-sparing and "masking" effects of salmeterol versus placebo in 13 asthmatic individuals requiring >= 1,500 microgram inhaled corticosteroid daily. Corticosteroid doses were reduced weekly until criteria were met for an exacerbation or the corticosteroid was fully withdrawn. Subjects were restabilized on their original dose of inhaled corticosteroid for 4 wk before crossover to the alternative treatment. Subjects maintained symptom and peak expiratory flow (PEF) diaries, and underwent weekly spirometric, methacholine challenge, sputum eosinophil, and serum eosinophil cationic protein (ECP) measurements. Mean corticosteroid dose was reduced by 87% during salmeterol treatment, versus 69% with placebo (p = 0.04). Sputum eosinophils increased before exacerbation despite stable symptoms, FEV1, and PEF. In the week before clinical exacerbation, sputum eosinophil counts were higher in the salmeterol-treatment arm (19.9 +/- 29.8% [mean +/- SD], versus placebo 9.3 +/- 17.6%; p = 0.006). Five subjects showed > 10% sputum eosinophilia before exacerbation during salmeterol treatment, as compared with two receiving placebo. In this model, salmeterol controlled symptoms and lung function until inflammation became significantly more advanced. We conclude that the bronchodilating and symptom-relieving effects of salmeterol can mask increasing inflammation and delay awareness of worsening asthma.     

Replication-deficient adenoviral vector for gene transfer potentiates airway neurogenic inflammation

Human trials for the treatment of cystic fibrosis lung disease with adenoviral vectors have been complicated by acute inflammatory reactions of unknown etiology. Because replicating respiratory viruses can potentiate tachykinin-mediated neurogenic inflammatory responses in airways, we studied whether the endotracheal administration of a replication-deficient adenoviral vector potentiated this response. The vector Ad5CMVLacZ was administered endotracheally to rats and the leakage of Evans blue dye was used to measure the capsaicin-induced neurogenic albumin extravasation. These studies show that neurogenic albumin extravasation is significantly potentiated in the airways of rats after administration of Ad5CMVLacZ. This inflammatory response can be blocked by selective antagonists of the substance P receptor or by glucocorticoids. Therefore, (1) the acute airway inflammation observed in patients after exposure to adenoviral vectors may exhibit a neurogenic component, which can be blocked pharmacologically, and (2) preclinical adenoviral vector safety studies of other organs innervated by the tachykinin system, e.g., coronary arteries and gastrointestinal tract, should include assessment of neurogenic inflammation.     

Use of induced sputum to examine airway inflammation in childhood asthma

Airway inflammation is important in the pathogenesis of asthma, during which it may lead to symptomatic exacerbations and increases in asthma severity, as well as contribute to future decline in asthma status. The use of induced sputum has emerged as an important and useful technique to study airway inflammation. It has particular advantages in the study of childhood asthma because it is noninvasive and allows samples to be collected on repeated occasions in children over 7 years of age. The results of cell counts are reliable when the sputum is processed in a standardized manner involving selection from saliva, cell dispersion, and quantitative cytology. Children with asthma have increased eosinophils and mast cells, which may persist even with high doses of inhaled corticosteroid therapy. During a severe exacerbation of asthma, there is an intense and heterogeneous inflammatory response involving eosinophil and neutrophil accumulation and activation. Characterization of the relevance of airway inflammation in children with asthma is important.     

Asthma and airway inflammation: potential anti-inflammatory activities of phosphodiesterase inhibitors

Although mental imagery has historically been ignored by the scientific community, it is one of the six most commonly used alternative therapies chosen by cancer patients. Investigations of immune response to psychosocial interventions have been largely anecdotal or quasi-experimental. Although reports of increased survival predominate among cancer patients, few investigators have measured psychological or immunological outcomes to explain the mechanisms that contribute to improved survival. The efficacy of imagery or support groups in patient adaptation (emotionally and physically) to the stresses of cancer, heightened immune function, and improved morbidity and mortality remain uncertain in the absence of objective evidence from randomized, controlled clinical trials. The imagery and group emotional support study pilot (IMAGES-P) is a 12-month feasibility study designed to examine the effect of group support and imagery/relaxation in a randomized, controlled clinical trial, differentiating the effects of these therapy modalities on immune function, quality of life, and the emotional well-being of women who have completed treatment for breast cancer. Secondary aims are to explore the quantitative relationships among emotional well-being, quality of life, and immune function. For this study established clinical trial methodology was used to examine functional immune responses associated with emotional states induced by the therapy interventions. As a pilot study, IMAGES-P will provide direction for future longitudinal studies evaluating the effectiveness of these interventions on emotional well-being and survival.     

Effects of airway inflammation on cough response in the guinea pig

We have developed a guinea pig model for cough related to allergic airway inflammation. Unanesthetized animals were exposed to capsaicin aerosols for 10 min, and cough frequency was counted during this period. The cough evaluation was performed by the following three methods: visual observation, acoustic analysis, and monitoring of pressure changes in the body chamber. These analyses clearly differentiated a cough from a sneeze. To elucidate the relationship between cough response and airway inflammation, animals were immunosensitized and multiple challenged. Sensitized guinea pigs presented no specific changes microscopically, but multiple-challenged animals showed an increased infiltration of inflammatory cells into the airway. Cough number in response to capsaicin increased significantly from 4.7 +/- 1.4 coughs/10 min in normal animals to 10.6 +/- 2.0 coughs/10 min in sensitized animals and further to 22.8 +/- 1.3 coughs/10 min in multiple-challenged animals. This augmented cough frequency was significantly inhibited by the inhalation of tachykinin-receptor antagonists and by oral ingestion, but not inhalation, of codeine phosphate. The results suggest that airway inflammation potentiates an elevation of cough sensitivity in this model.     

Differential recruitment of leukocyte populations and alteration of airway hyperreactivity by C-C family chemokines in allergic airway inflammation

Allergic airway inflammation is characterized by peribronchial leukocyte accumulation within the airway. Subsequent tissue damage leading to airway hyperreactivity is a result of activation of multiple leukocyte populations. Using an established model of allergic airway inflammation induced by intratracheal challenge with parasite (Schistosoma mansoni) egg Ag in presensitized mice, we have examined differential leukocyte recruitment. These studies have identified key chemokines involved in the accumulation of specific subsets of cells and the induction of airway hyperreactivity. In this study we have examined three C-C family chemokines, MCP-1, MIP-1alpha, and RANTES, which promote mononuclear cell- and eosinophil-specific recruitment to the airway. The in vivo neutralization of either MIP-1alpha or RANTES, but not MCP-1, significantly reduced the intensity of the eosinophil recruitment to the lung and airway during the allergic airway response by >50 and >60%, respectively. In contrast, neutralization of MCP-1 significantly reduced total leukocyte migration (>50% reduction), whereas neutralization of RANTES and MIP-1alpha had no significant affect on the overall leukocyte migration. Further examination of the effect of MCP-1 depletion indicated that both CD4+ and CD8+ lymphocyte subsets were decreased. Depletion of MCP-1 significantly reduced the airway hyperreactivity to near control levels, whereas depletion of MIP-1alpha or RANTES did not affect the intensity of airway hyperreactivity. These data indicate that multiple C-C chemokines are involved in the recruitment of particular leukocyte populations and that neutralization of MCP-1, but not RANTES or MIP-1alpha, significantly reduced airway hyperreactivity.     

Natural killer cells determine development of allergen-induced eosinophilic airway inflammation in mice

The earliest contact between antigen and the innate immune system is thought to direct the subsequent antigen-specific T cell response. We hypothesized that cells of the innate immune system, such as natural killer (NK) cells, NK1.1(+) T cells (NKT cells), and gamma/delta T cells, may regulate the development of allergic airway disease. We demonstrate here that depletion of NK1.1(+) cells (NK cells and NKT cells) before immunization inhibits pulmonary eosinophil and CD3(+) T cell infiltration as well as increased levels of interleukin (IL)-4, IL-5, and IL-12 in bronchoalveolar lavage fluid in a murine model of allergic asthma. Moreover, systemic allergen-specific immunoglobulin (Ig)E and IgG2a levels and the number of IL-4 and interferon gamma-producing splenic cells were diminished in mice depleted of NK1.1(+) cells before the priming regime. Depletion of NK1.1(+) cells during the challenge period only did not influence pulmonary eosinophilic inflammation. CD1d1 mutant mice, deficient in NKT cells but with normal NK cells, developed lung tissue eosinophilia and allergen-specific IgE levels not different from those observed in wild-type mice. Mice deficient in gamma/delta T cells showed a mild attenuation of lung tissue eosinophilia in this model. Taken together, these findings suggest a critical role of NK cells, but not of NKT cells, for the development of allergen-induced airway inflammation, and that this effect of NK cells is exerted during the immunization. If translatable to humans, these data suggest that NK cells may be critically important for deciding whether allergic eosinophilic airway disease will develop. These observations are also compatible with a pathogenic role for the increased NK cell activity observed in human asthma.     

Repeated subacute ozone exposure of inbred mice: airway inflammation and ventilation

The present study was designed to assess the effects of repeated subacute ozone (O3) exposure on pulmonary inflammation and ventilation in two inbred strains of mice differentially susceptible to a single O3 exposure. Susceptible C57BL/6J (B6) and resistant C3H/HeJ (C3) mice were exposed to 0.3 ppm O3 for 48 and 72 h and, after 14 days recovery, both strains were reexposed. Airway inflammation and lung injury were assessed by counting inflammatory cells and measuring total protein content and lactate dehydrogenase (LDH) activity in bronchoalveolar lavage (BAL) returns. Minute ventilation [VE, the product of breathing frequency (f), and tidal volume (VT)] was measured prior to and immediately following each exposure. After the initial exposure, B6 mice developed greater O3-induced increases in total protein, inflammatory cell influx, and LDH activity compared to C3 mice. In normal air, VE was also significantly elevated in B6, but not C3, mice after O3. The hypercapnic f of B6 and hypercapnic VT of C3 mice were significantly altered after O3 exposure. Reexposure to O3 caused a smaller increase in the numbers of macrophages, lymphocytes, epithelial cells, and BAL protein in both strains, and no changes in LDH activity. However, the number of polymorphonuclear leukocytes significantly increased in B6 and C3 mice as compared to the initial O3 exposure. In both strains, the ventilatory responses to normal air or hypercapnia were largely reproducible after O3 reexposure. Results indicated that differential susceptibility to O3-induced inflammation was maintained in B6 and C3 mice with O3 reexposure although the magnitude of the difference was reduced. Results also suggest that the ventilatory responses to O3 in B6 and C3 mice were reproducible with reexposure, and that airway inflammation and ventilation were not codependent.     

Severity of airflow limitation is associated with severity of airway inflammation in smokers

To investigate the relationship between airflow limitation and airway inflammation in smokers, we examined paraffin-embedded bronchial biopsies obtained from 30 smokers: 10 with severe airflow limitation, eight with mild/moderate airflow limitation, and 12 control smokers with normal lung function. Histochemical and immunohistochemical methods were performed to assess the number of inflammatory cells in the subepithelium and the expression of CC chemokines macrophage inflammatory protein (MIP)-1alpha and -1beta in the bronchial mucosa. Compared with control smokers, smokers with severe airflow limitation had an increased number of neutrophils (p < 0.02), macrophages (p < 0.03), and NK lymphocytes (p < 0.03) in the subepithelium, and an increased number of MIP-1alpha+ epithelial cells (p < 0.02). When all smokers were considered together, the value of FEV1 was inversely correlated with the number of neutrophils (r = -0.59, p < 0.002), macrophages (r = -047, p < 0. 012), NK-lymphocytes (r = -0.51, p < 0.006) in the subepithelium, and with the number of MIP-1alpha+ epithelial cells (r = -0.61, p < 0.003). We conclude that in smokers the severity of airflow limitation is correlated with the severity of airway inflammation and that severe airflow limitation is associated with an increased number of neutrophils, macrophages, NK lymphocytes, and MIP-1alpha+ cells in the bronchial mucosa.