Phenotyping of difficult asthma using longitudinal physiological and biomarker measurements reveals significant differences in stability between clusters.

BACKGROUND: Although the heterogeneous nature of asthma has prompted asthma phenotyping with physiological or biomarker data, these studies have been mostly cross-sectional. Longitudinal studies that assess the stability of phenotypes based on a combination of physiological, clinical and biomarker data are currently lacking. Our objective was to assess the longitudinal stability of clusters derived from repeated measures of airway and physiological data over a 1-year period in moderate and severe asthmatics. METHODS: A total of 125 subjects, 48 with moderate asthma (MA) and 77 with severe asthma (SA) were evaluated every 3 months and monthly, respectively, over a 1-year period. At each 3-month time point, subjects were grouped into 4 asthma clusters (A, B, C, D) based on a combination of clinical (duration of asthma), physiological (FEV1 and BMI) and biomarker (sputum eosinophil count) variables, using k-means clustering. RESULTS: Majority of subjects in clusters A and C had severe asthma (93 % of subjects in cluster A and 79.5 % of subjects in cluster C at baseline). Overall, a total of 59 subjects (47 %) had stable cluster membership, remaining in clusters with the same subjects at each evaluation time. Cluster A was the least stable (21 % stability) and cluster B was the most stable cluster (71 % stability). Cluster stability was not influenced by changes in the dosage of inhaled corticosteroids. CONCLUSION: Asthma phenotyping based on clinical, physiologic and biomarker data identified clusters with significant differences in longitudinal stability over a 1-year period. This finding indicates that the majority of patients within stable clusters can be phenotyped with reasonable accuracy after a single measurement of lung function and sputum eosinophilia, while patients in unstable clusters will require more frequent evaluation of these variables to be properly characterized.

Induction of GITRL expression in human keratinocytes by Th2 cytokines and TNF-α: implications for atopic dermatitis.

BACKGROUND: Glucocorticoid-induced TNF receptor-related protein ligand (GITRL), a ligand for the T cell co-stimulatory molecule GITR, is expressed by keratinocytes and involved in chemokine production. The expression of GITRL in skin inflammation remains unknown. OBJECTIVES: This study investigated cytokine regulation of keratinocyte GITRL expression. METHODS: Glucocorticoid-induced TNF receptor expression was evaluated in cytokine-treated human epidermal keratinocytes (HEK)s, murine PAM 212 cell line, murine and human skin explants by real time PCR, flow cytometry and immunostaining. Functional responses to GITR fusion protein were examined by real time PCR and ELISA. GITRL expression in AD and psoriasis was studied by immunohistochemistry. RESULTS: Skin biopsies from STAT6VT transgenic mice, which develop spontaneous atopic skin inflammation, were found by immunofluoresence, to have increased keratinocyte GITRL expression. Exposure to Th2 cytokines augmented GITRL mRNA expression in the murine PAM 212 keratinocytic cell line and murine skin explants. In contrast, GITRL mRNA and protein expression was only increased in HEKs and human skin explants in the presence of the combination of TNF-α and Th2 cytokines. A synergistic effect of Th2 cytokines and GITR fusion protein on production of CCL17, the Th2 chemokine, by murine keratinocytes was demonstrated. Immunohistochemical staining showed that acute AD lesions have increased expression of GITRL compared with normal skin, chronic AD lesions and psoriatic plaques. CONCLUSIONS AND CLINICAL RELEVANCE: Our studies demonstrate that GITRL expression is augmented by Th2 cytokines and TNF-α in keratinocytes. Increased GITRL expression in acute AD skin lesions is shown. This observation suggests a link between cytokine-regulated keratinocyte GITRL expression and its role in inflammatory responses in AD.

Conjugates of ovalbumin and monomethoxypolyethylene glycol abolish late allergic responses and decrease IL-4 and IL-5 mRNA expression in the rat.

The purpose of this study was to test the therapeutic potential of monomethoxypolyethylene glycol (mPEG) conjugated-allergen using a rodent model of allergic asthma. Previously, this conjugate has been shown to possess the dual capacity of inducing long-term ovalbumin (OA)-specific suppression of the antibody response and inactivating rat mast cells that have been sensitized with murine IgE to OA. Ovalbumin sensitized and challenged Brown Norway rats were studied. Fourteen days after sensitization, a test group of six rats received mPEG-OA solution intratracheally and were challenged 30 min later with aerosolized OA. Another group of seven sensitized rats was similarly challenged with OA 30 min after intratracheal administration of normal saline. A group of six sensitized rats received mPEG-OA solution intratracheally but were challenged with normal saline. Another group of seven sensitized rats received mPEG-BSA solution intratracheally and were challenged 30 min later with aerosolized OA. A final group of five unsensitized rats were neither challenged nor medicated intratracheally. Pulmonary resistance was measured before and for 8 h following inhalation challenge. mPEG-OA treatment had an inhibitory effect on the allergic late airway response, but the early response was not significantly altered. Both mPEG-OA and mPEG-BSA reduced the total cells, eosinophils and neutrophils, in bronchoalveolar lavage and decreased the expression of IL-4, IL-5 and IFN-gamma mRNA. In conclusion, mPEG-OA can prevent the development of allergen-induced late airway responses and reduce airway Th2-type cytokine expression whereas mPEG conjugated to an irrelevant antigen (BSA) is anti-inflammatory but does not affect the late response.

Neutrophilic airway inflammation in horses with heaves is characterized by a Th2-type cytokine profile.

Heaves in horses shares many similarities with human asthma, including lower airway inflammation, reversible airway obstruction, and bronchial hyperresponsiveness. Extrinsic asthma is an allergic response to environmental allergens and a similar immunologic mechanism may be implicated in heaves. It is now recognized that a Th2 subset of CD4+ lymphocytes is associated with allergic diseases such as atopic asthma. The purpose of this study was to determine whether airway inflammation in heaves is associated with a pattern of expression of cytokine suggestive of a Th2 type response. The expression of mRNA, encoding interleukin (IL)-4, IL-5, and interferon gamma (IFN-gamma) was measured in bronchoalveolar cells from seven horses with heaves and five control horses, using in situ hybridization and radiolabeled equine-specific cRNA probes coding for these cytokines. Bronchoalveolar cells of horses with heaves had an increased expression of IL-4 (p = 0.01) and IL-5 (p = 0.02) mRNA and a decreased expression of INF-gamma (p = 0.01) compared with control horses. Here we show that inflammatory cells in lungs from horses with heaves display a Th2-type cytokine profile that is consistent with the hypothesis that heaves is an allergic condition with similarity to human asthma.

GRbeta expression in nasal polyp inflammatory cells and its relationship to the anti-inflammatory effects of intranasal fluticasone.

BACKGROUND: Nasal polyposis disease is an inflammatory disorder with intense eosinophilic infiltration of respiratory mucosa that is often difficult to control with topical steroids. Recent evidence suggests that overexpression of the glucocorticoid receptor splice variant GRbeta in inflammatory cells might contribute to steroid insensitivity in diseases such as asthma. OBJECTIVE: The purposes of this investigation were to determine whether nasal polyp (NP) inflammatory cells overexpress GRbeta and to examine whether GRbeta overexpression is associated with insensitivity to the potent topical steroid fluticasone propionate (FP). METHODS: Biopsies were obtained from 10 subjects with NPs before and 4 weeks after treatment with intranasal FP. Middle turbinates biopsies from 6 healthy, nonallergic subjects served as normal controls. Biopsies were immunostained for inflammatory cell markers as well as GRbeta and probed for various cytokine mRNA. The anti-inflammatory response to FP was examined in relation to pretreatment levels of GRbeta expression. RESULTS: The total numbers of inflammatory cells were increased in NPs. The percentage of inflammatory cells expressing GRbeta was also increased (40.5% +/- 19.2% vs 16.1% +/- 4.0%, P =.009). GRbeta expression in NPs was almost exclusive to T lymphocytes, eosinophils, and macrophages. An inverse correlation was observed between the baseline inflammatory cell GRbeta expression and the reduction after FP treatment in EG2-positive eosinophils, CD4-positive T lymphocytes, endothelial VCAM-1 expression, and IL-4 mRNA-positive cells. NPs that were "FP-insensitive" in terms of suppression of eosinophil numbers (major basic protein-positive) had a significantly greater percentage of GRbeta-positive inflammatory cells, a higher ratio of GRbeta-positive/GRalpha-positive cells, and increased numbers of GRbeta-positive eosinophils and macrophages in comparison with those that were "FP-sensitive." "FP-insensitive" NPs also demonstrated a higher percentage of IL-5-positive inflammatory cells expressing GRbeta before and after FP treatment. CONCLUSION: GRbeta expression appears to be a marker of steroid insensitivity in NPs. Expression of GRbeta by NP inflammatory cells, particularly T cells and eosinophils, might render them resistant to suppression by topical steroids and thereby contribute to persistent NP inflammation.

Anti-inflammatory activity of clarithromycin in adults with chronically inflamed sinus mucosa.

In a phase IV, open-label study, 25 patients with clinically stable chronic sinusitis and persistent maxillary sinus inflammation were treated for 14 days with clarithromycin 500 mg twice daily. Biopsy specimens of the maxillary sinus mucosa were obtained pretreatment and evaluated for macrophages (CD68), eosinophils (MBP), elastase, interleukin-6 (IL-6), IL-8, tumor necrosis factor-alpha (TNF-alpha), and activity of eosinophils (EG2), as well as edema score. Clinical signs and symptoms were assessed pretreatment, at the end of treatment, and 1 and 2 weeks later. Statistically significant reductions (P < or = .05) from pretreatment were observed for all markers of sinus mucosal inflammation, including CD68, EG2, elastase, IL-6, IL-8, TNF-alpha, and edema score, with a trend to decreased total eosinophil count. Improvement was observed for all clinical signs and symptoms of chronic sinusitis--sinus pain, sinus headache, nasal congestion, nasal discharge, and mucopurulent discharge--up to 14 days after the end of treatment. Cultures to evaluate persistent infection with Chlamydia pneumoniae showed negative results. Significant reductions in various markers of sinus mucosal inflammation support the role of clarithromycin in modulating immunologic responses. Improvement of clinical signs and symptoms in patients with chronic inflammatory sinusitis not meeting criteria for known or presumed bacterial infection was also noted up to 2 weeks after completion of a 14-day course of clarithromycin.

Gene expression of vascular endothelial growth factor and its receptors and angiogenesis in bronchial asthma.

BACKGROUND: Angiogenesis is a feature of airway remodeling in bronchial asthma. The mechanism responsible for this angiogenesis is unknown. Vascular endothelial growth factor (VEGF) is a potent inducer of endothelial cells, which may contribute to chronic inflammation and angiogenesis. OBJECTIVE: We sought to investigate the molecular mechanisms underlying increased vascularity, and we examined the mRNA expression of VEGF and its receptors (flt-1 and flk-1) within bronchial biopsy specimens from asthmatic patients and normal control subjects. METHODS: Endobronchial biopsy specimens were examined immunocytochemically by staining with anti-type IV collagen mAb to evaluate vessel density by using computer-assisted image analysis. Specimens were also analyzed for the presence of the mRNAs of VEGF and its receptors with in situ hybridization. RESULTS: The extent of airway vascularity was increased in asthmatic subjects compared with that in control subjects (P <.01). Asthmatic subjects exhibited a greater expression of VEGF, flt-1, and flk-1 mRNA(+) cells in the airway mucosa compared with that in control subjects (P <.001 for each comparison). The degree of vascularity was associated with the number of VEGF, flt-1, and flk-1 mRNA(+) cells. Numbers of cells expressing VEGF mRNA inversely correlated with airway caliber (r = -0.83, P <.01) and airway hyperresponsiveness (r = -0.97, P <.001). Colocalization studies showed that macrophages, eosinophils, and CD34(+) cells were the major sources of VEGF; CD34(+) cells, macrophages, and T cells expressed both flt-1 and flk-1. CONCLUSION: These findings provide evidence that VEGF may play an important role in angiogenesis and subsequent airway remodeling in bronchial asthma.

Inflammatory patterns of allergic and nonallergic rhinitis.

Rhinitis is a chronic condition of the nasal mucosa that affects a large segment of the population. The symptoms of rhinitis occur in a variety of sinonasal conditions, which may be broadly classified as allergic (seasonal or perennial) or nonallergic (infectious or a number of noninfectious etiologies) based on the presence or absence of atopy. The cytokine profile and inflammatory patterns underlying these two conditions vary because of certain differences in their pathophysiology as discussed in this review.

Recruitment of T cells to the lung in response to antigen challenge.

T-cell recruitment to the lungs is thought to represent a key step in airway allergic inflammation. T cells coordinate and amplify effector functions of antigen-specific and nonspecific proinflammatory cells, such as B cells and eosinophils. The T(H)2 cell, in particular, promotes allergic inflammation through the expression of IL-4, IL-5, and IL-13, proinflammatory cytokines that are important in the induction of B-cell switching and the promotion of eosinophil proliferation and survival. This cytokine profile has been implicated in asthma; elevations in bronchoalveolar lavage IL-4 and IL-5 levels have been observed in asthmatic patients. The recruitment of T(H) cells to the site of allergic inflammation (lung) is the subject of this review.

Regulation of IL-1-induced gingival collagenase gene expression by activator protein-1 (c-Fos/c-Jun).

Matrix metalloproteinase-1 is probably involved in the progression of periodontal disease. The aim of this study was to investigate whether IL-1beta stimulates the expression of the activator protein 1 (AP-1) transcription factor and, consequently, if the AP-1 transcription factor participates in the regulation of collagenase gene expression in human gingival fibroblast cells. In this study, we demonstrate that the concentration of the protein components of AP-1 transcription factor, c-Fos and c-Jun, is enhanced by IL-1beta both at mRNA and protein levels, utilizing Northern blot analysis, electrophoretic mobility gel shift assay and Western blot analysis. The IL-1beta stimulated the collagenase-CAT and AP-1-CAT activities in a dose dependent manner with respect to the amount of DNA used in transfections. Further, overexpression of c-Fos and c-Jun proteins revealed a dose-dependent transcriptional activation of the collagenase promoter. These findings, coupled with the existence of AP-1 consensus DNA binding sites on the collagenase gene promoter, show that regulation of collagenase gene expression by IL-1beta involves the transcription factor AP-1 in gingival fibroblasts.

Increased number of glucocorticoid receptor-beta-expressing cells in the airways in fatal asthma.

BACKGROUND: We have recently demonstrated an increased number of glucocorticoid receptor-beta (GRbeta)-positive cells in steroid-insensitive subjects with severe asthma. Insensitivity to steroids may be a major contributing factor in fatal asthma; however, no such direct evidence has been report previously. OBJECTIVE: Our aims were to investigate the expression of GRbeta immunoreactivity, an endogenous inhibitor of steroid action previously associated with steroid insensitivity, within the airways of patients who died of slow-onset fatal asthma and to compare its expression in patients with emphysema and in nonasthmatic subjects who died of unrelated causes. Sections from airways, both large and small, were obtained from 7 patients who died of asthma, 6 who died from emphysema, and 8 who died from nonpulmonary diseases. Sections from lungs of 6 patients with mild asthma whose lungs were resected for carcinoma were also included as controls. METHODS: Tissue samples were processed for immunocytochemistry with a polyclonal antibody to GRbeta with use of the avidin-biotin technique and with monoclonal CD3, major basic protein, CD68, and elastase antibodies with the alkaline phosphatase-anti-alkaline phosphatase technique. Sequential immunocytochemistry was performed to phenotype the GRbeta immunoreactive cells. Tissue sections from both large (>2 mm) and small (<2 mm) airways were examined. RESULTS: There was a significantly greater number of GRbeta immunoreactive cells in fatal asthma compared with emphysema and controls (P <.001 and P <.05, respectively). There was no difference in the expression of GRbeta in emphysema compared with controls. GRbeta immunoreactivity was also significantly higher in fatal asthma compared with mild asthma. The expression of GRbeta in the small airways of patients with severe asthma did not differ significantly from that in the large airways. The majority of GRbeta-positive cells were T cells and to a lesser extent eosinophils, macrophages, and neutrophils. CONCLUSION: The results of this study support the association of GRbeta expression with fatal asthma and suggest that alternative anti-inflammatory agents need to be considered in the acute setting for patients who are not responding to steroid therapy.

The pathology of chronic asthma.

Our understanding of the pathophysiology of asthma has undergone great advances in the past decade, particularly with the recognition of cytokines and the roles they may take in orchestrating the local immune response. With this information, it has been possible to target new therapeutic entities such as cytokine or chemokine receptors. Eosinophils and T lymphocytes have a special place in the inflammatory and structural alterations contributing to the asthmatic diathesis. It is possible that phenotype subsets of these cells exist and they hold the key to perpetuation of immunologic and physiologic abnormalities in asthma.

Evidence for increased expression of eotaxin and monocyte chemotactic protein-4 in atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease associated with tissue eosinophilia and the activation of T lymphocytes. The novel eosinophil chemoattractants, eotaxin and monocyte chemotactic protein (MCP)-4, are up-regulated at sites of allergic inflammation, yet their contribution to the pathophysiologic mechanisms of AD remains to be determined. OBJECTIVE: We sought to investigate the expression of eotaxin and MCP-4 in acute and chronic lesions from patients with AD and to determine their relationship to the numbers of resident inflammatory cells. METHODS: With use of in situ hybridization, the expression of eotaxin and MCP-4 messenger RNA (mRNA) in skin biopsy specimens from patients with acute and chronic AD skin lesions was compared with that of uninvolved skin from these patients and skin from healthy volunteers. RESULTS: There was a constitutive expression of eotaxin and MCP-4 mRNA in skin biopsy specimens from healthy subjects. Positive signal for chemokine mRNA was observed both within the epidermis and inflammatory cells (macrophages, eosinophils, and T cells) of the subepidermis in AD skin lesions. Within the subepithelium acute and chronic skin lesions exhibited a significant increase in the numbers of eotaxin and MCP-4 mRNA-positive cells compared with uninvolved skin (P <.01), whereas the numbers of eotaxin and MCP-4 mRNA-positive cells were significantly higher in chronic AD compared with acute AD skin lesions (P <.005, P <.001, respectively). Correlations were observed between the expression of eotaxin and MCP-4 mRNA and the presence of eosinophils and macrophages, respectively, in AD lesions (r(2) = 0.84, r(2) = 0.94). CONCLUSION: There is an increased expression of eotaxin and MCP-4 in acute and chronic lesions, suggesting that these chemotactic factors play a major role in the pathophysiologic mechanisms of AD.

Induction of corticosteroid insensitivity in human PBMCs by microbial superantigens.

BACKGROUND: Microbial superantigens have been described to contribute to the pathogenesis of chronic inflammatory diseases often complicated by insensitivity to glucocorticoid therapy. In bronchial asthma glucocorticoid insensitivity has been associated with increased expression of glucocorticoid receptor beta, an endogenous inhibitor of the classic glucocorticoid receptor alpha. OBJECTIVE: To study a potential mechanism by which superantigens could contribute to poor disease control, we examined their capacity to alter steroid sensitivity and expression of glucocorticoid receptor beta. METHODS: The capacity of dexamethasone to inhibit stimulation of PBMCs from 7 healthy subjects with the prototypic superantigens, staphylococcal enterotoxin (SE) B, toxic shock syndrome toxin (TSST)-1 and SEE, versus PHA, was tested. The expression of glucocorticoid receptor beta in normal PBMCs after stimulation with SEB, versus PHA, was assessed by immunocytochemistry. RESULTS: Dexamethasone 10(-6) mol/L caused a 99% inhibition of PHA-induced PBMC proliferation but only a 19% inhibition of the SEB-induced, 26% inhibition of the TSST-1, and 29% inhibition of the SEE-induced PBMC proliferation (P <.01 for all superantigens versus PHA) demonstrating that superantigens can induce steroid insensitivity. Stimulation of normal PBMCs with SEB induced a significant increase of glucocorticoid receptor beta compared with PHA and unstimulated cells (P <.01). CONCLUSION: We have demonstrated the capacity of microbial superantigens to induce glucocorticoid insensitivity, which should be considered in the diagnosis and treatment of patients with superantigen-triggered diseases. These data suggest that superantigens may contribute to glucocorticoid insensitivity through induction of glucocorticoid receptor beta.

Molecular pathology of allergic disease: I: lower airway disease.

Asthma is a complex disorder associated with eosinophil infiltration and the activation of T lymphocytes within the airways. Recent advances in the pathophysiologic mechanisms of asthma point to the importance of eosinophil-basophil progenitor cells and a family of transcription factors that underlie the development of T(H)2-type responses. Further research is needed to address the development of chronic inflammatory changes, the role of profibrotic cytokines, and especially their reliance on eosinophils in the lungs.

IL-9 and its receptor in allergic and nonallergic lung disease: increased expression in asthma.

BACKGROUND: Bronchial asthma is a chronic inflammatory disease associated with genetic components. Recently IL-9 has been reported as a candidate gene for asthma and to be associated with bronchial hyperresponsiveness and elevated levels of total serum IgE. OBJECTIVE: To investigate the contribution of IL-9 to the pathogenesis of asthma, we examined the expression of IL-9 and its receptor (IL-9R) in bronchial tissue from subjects with atopic asthma (n = 10), chronic bronchitis (n = 11), and sarcoidosis (n = 9) and from atopic (n = 7) and nonatopic (n = 10) healthy control subjects. METHODS: Bronchial biopsy specimens were examined for the presence of IL-9 and IL-9R protein and messenger RNA (mRNA) by immunocytochemistry and in situ hybridization, respectively. To phenotype the cells expressing IL-9 in asthmatic tissue, combined in situ hybridization and immunocytochemistry was also performed. RESULTS: There was a highly significant difference (P <.001) in the expression of IL-9 mRNA in asthmatic airways (20.6 +/- 4.0 cells/mm of basement membrane) compared with chronic bronchitis (5.6 +/- 4.4), sarcoidosis (2.5 +/- 1.8), atopic control subjects (7.7 +/- 2.2), and healthy control subjects (2.7 +/- 2.3). The number of IL-9 immunoreactive cells was also greater in asthmatic patients compared with the other groups (P <.05). Although the level of IL-9R mRNA expression did not differ in any of the groups (P >.05), IL-9R immunoreactivity was significantly higher in asthmatic compared with control subjects. Furthermore, IL-9 mRNA expression levels were also significantly correlated with FEV(1) (P <.05) and the airway responsiveness to methacholine producing a 20% fall in FEV(1) (P <. 01). The cells expressing IL-9 mRNA in asthmatic tissue were CD3(+) lymphocytes (68%), major basic protein(+) eosinophils (16%), and elastase(+) neutrophils (8%). CONCLUSION: The results of this study demonstrate the potential of IL-9 to be a marker for atopic asthma and furthermore suggest an important role for this cytokine in the pathophysiologic mechanisms of this disease.

Epidermal HLA-DR and the enhancement of cutaneous reactivity to superantigenic toxins in psoriasis.

Streptococcal and staphylococcal superantigens (SAg's) have been implicated in the pathogenesis of inflammatory skin diseases, but the mechanisms by which these toxins act are unknown. The present study assessed the ability of nanogram quantities of topically applied purified toxic shock syndrome toxin-1 (TSST-1), staphylococcal enterotoxin type B, and streptococcal pyrogenic enterotoxin types A and C to induce inflammatory reactions in clinically uninvolved skin of normal controls and subjects with psoriasis, atopic dermatitis, and lichen planus. These SAg's triggered a significantly greater inflammatory skin response in psoriatics than in normal control subjects or in subjects with atopic dermatitis or lichen planus. Surprisingly, skin biopsies did not exhibit the T-cell receptor Vbeta stimulatory properties predicted for SAg-induced skin reactions. By 6 hours after patch testing with SAg's, TNF-alpha mRNA had increased in the epidermis (but not the dermis) in biopsies from psoriatics, compared with controls. Immunohistochemical studies revealed significantly higher HLA-DR expression in keratinocytes from psoriatics than from controls. However, a mutant TSST-1 protein that fails to bind HLA-DR did not elicit an inflammatory skin reaction. These results indicate that keratinocyte expression of HLA-DR enhances inflammatory skin responses to SAg's. They may also account for previous studies failing to demonstrate selective expansion of T-cell receptor Vbetas in psoriatics colonized with SAg-producing Staphylococcus aureus, and they identify a novel T cell-independent mechanism by which SAg's contribute to the pathogenesis of inflammatory skin diseases.

Increased expression of IL-12 receptor mRNA in active pulmonary tuberculosis and sarcoidosis.

Cytokines have been implicated in the pathophysiology and development of pulmonary diseases such as tuberculosis and sarcoidosis. In particular, the numbers of cells expressing Th1-type cytokines such as IFN-gamma and IL-12 are increased within the lungs of patients with these granulomatous diseases. As a factor promoting the commitment of naive lymphocytes to a Th1-type profile of cytokine expression, IL-12 may be pivotal in the cascade of proinflammatory events within the airways. In this study, we examined the expression of the IL-12 receptor (IL-12R) mRNA in bronchoalveolar lavage (BAL) fluid from patients with active pulmonary tuberculosis (n = 6) and active pulmonary sarcoidosis (n = 6), and from allergic asthmatics (n = 6) and normal control subjects (n = 6). Bronchoscopy with BAL was undertaken, and cell cytospins were examined using the technique of in situ hybridization. There was a significant increase in the numbers of cells expressing mRNA for both beta(1) and beta(2) subunits of the IL-12R in active pulmonary sarcoidosis (p < 0.02, p < 0.01, respectively) and active pulmonary tuberculosis (p < 0.01, p < 0.005, respectively) compared with normal control subjects. In contrast, the allergic asthmatic patients exhibited a significant decrease in the number of IL-12R mRNA-positive cells (both beta(1) and beta(2) subunits (p < 0.01, p < 0.005, respectively), compared with the normal control subjects. These patients did, however, exhibit a significant increase in IL-4R mRNA, which was not evident in those with either tuberculosis or sarcoidosis when compared with normal subjects (p < 0.05). Colocalization studies demonstrated that CD8+ve cells are a principal site for the expression of IL-12R in tuberculosis. In sarcoidosis, IL-12R was expressed both on CD4+ve and CD8+ve cells. The increased expression of receptors for IL-12 in granulomatous diseases such as pulmonary tuberculosis and sarcoidosis provides evidence supporting the commitment of lymphocytes to a Th1-type cytokine profile in vivo.

Airway epithelium expresses interleukin-18.

Interleukin (IL)-18 is an interferon (IFN)-gamma-inducing cytokine suggested to be important in regulating inflammatory responses. This study investigated the pulmonary expression of IL-18 under conditions characterized by T-helper (Th)1 (lipopolysaccharide (LPS) treatment/sarcoidosis) and Th2 (ovalbumin (OVA) challenge/asthma) cytokine production. In situ hybridization and immunocytochemistry were used to determine the number of cells expressing IL-18, IFN-gamma, IL-5 and major basic protein (MBP) within lung tissue from Balb/c mice stimulated with LPS, OVA and in normal control mice. Bronchial biopsies from patients with sarcoidosis, asthma and control individuals were also examined. IL-18 was localized primarily to airway epithelium and mononuclear cells. Constitutive expression was observed within the lungs of control mice. Animals challenged with LPS exhibited more IL-18 messenger ribonucleic acid (mRNA)-positive and IFN-gamma immunoreactive cells, compared to control mice (p<0.01). OVA-challenged mice had fewer IL-18 mRNA positive and more IL-5 and MBP immunoreactive cells, compared to control mice (p<0.01). Similarly, constitutive expression of IL-18 protein was observed within the airway epithelium of control individuals, with more positive cells found within sarcoidosis tissue (p<0.01) and fewer within asthmatic tissue (p<0.01), compared to controls. These results demonstrate the expression of interleukin-18 within airway epithelium and the regulation of this cytokine under conditions of both T-helper1 and T-helper2 cytokine production.