Challenging immunodominance of influenza-specific CD8+ T cell responses restricted by the risk-associated HLA-A*68:01 allomorph.

Although influenza viruses lead to severe illness in high-risk populations, host genetic factors associated with severe disease are largely unknown. As the HLA-A*68:01 allele can be linked to severe pandemic 2009-H1N1 disease, we investigate a potential impairment of HLA-A*68:01-restricted CD8+ T cells to mount robust responses. We elucidate the HLA-A*68:01+CD8+ T cell response directed toward an extended influenza-derived nucleoprotein (NP) peptide and show that only ~35% individuals have immunodominant A68/NP145+CD8+ T cell responses. Dissecting A68/NP145+CD8+ T cells in low vs. medium/high responders reveals that high responding donors have A68/NP145+CD8+ memory T cells with clonally expanded TCRαßs, while low-responders display A68/NP145+CD8+ T cells with predominantly naïve phenotypes and non-expanded TCRαßs. Single-cell index sorting and TCRαß analyses link expansion of A68/NP145+CD8+ T cells to their memory potential. Our study demonstrates the immunodominance potential of influenza-specific CD8+ T cells presented by a risk HLA-A*68:01 molecule and advocates for priming CD8+ T cell compartments in HLA-A*68:01-expressing individuals for establishment of pre-existing protective memory T cell pools.

Human leucocyte antigen-defined microchimerism early post-transplant does not predict for stable lung allograft function.

Microchimerism is the presence of foreign cells in an individual below 1% of total cells, which can occur in the setting of solid organ transplantation. This study quantitated donor-derived cellular subsets longitudinally in human leucocyte antigen (HLA)-mismatched lung transplant recipients (LTR) during the first post-operative year and evaluated the pattern of peripheral microchimerism with clinical outcomes. Peripheral blood mononuclear cells (PBMC) isolated from non-HLA-B44 LTR who received HLA-B44 allografts were sorted flow cytometrically into three cellular subsets. Real-time quantitative polymerase chain reaction (q-PCR) demonstrated that donor-derived HLA-B44 microchimerism is a common phenomenon, observed in 61% of patients. The level of donor-derived cells varied across time and between LTR with frequencies of 38% in the B cells/monocytes subset, 56% in the T/NK cells subset and 11% in the dendritic cells (DC) subset. Observations highlighted that microchimerism was not necessarily associated with favourable clinical outcomes in the first year post-lung transplantation.

Cytomegalovirus replication within the lung allograft is associated with bronchiolitis obliterans syndrome.

Early studies reported cytomegalovirus (CMV) pneumonitis as a risk factor for development of bronchiolitis obliterans syndrome (BOS) following lung transplantation. While improvements in antiviral prophylaxis have resulted in a decreased incidence of CMV pneumonitis, molecular diagnostic techniques allow diagnosis of subclinical CMV replication in the allograft. We hypothesized that this subclinical CMV replication was associated with development of BOS. We retrospectively evaluated 192 lung transplant recipients (LTR) from a single center between 2001 and 2009. Quantitative (PCR) analysis of CMV viral load and histological evidence of CMV pneumonitis and acute cellular rejection was determined on 1749 bronchoalveolar lavage (BAL) specimens and 1536 transbronchial biopsies. CMV was detected in the BAL of 41% of LTR and was significantly associated with the development of BOS (HR 1.8 [1.1-2.8], p = 0.02). This association persisted when CMV was considered more accurately as a time-dependent variable (HR 2.1 [1.3-3.3], p = 0.003) and after adjustment for significant covariates in a multivariate model. CMV replication in the lung allograft is common following lung transplantation and is associated with increased risk of BOS. As antiviral prophylaxis adequately suppresses CMV longer prophylactic strategies may improve long-term outcome in lung transplantation.

Randomised placebo controlled trial of non-invasive ventilation for hypercapnia in cystic fibrosis.

BACKGROUND: The clinical benefits of domiciliary non-invasive positive pressure ventilation (NIV) have not been established in cystic fibrosis (CF). We studied the effects of nocturnal NIV on quality of life (QoL), functional and physiological outcomes in CF subjects with awake hypercapnia (arterial carbon dioxide pressure PaCO2>45 mm Hg). METHODS: In a randomised, placebo controlled, crossover study, eight subjects with CF, mean (SD) age 37 (8) years, body mass index 21.1 (2.6) kg/m2, forced expiratory volume in 1 s 35 (8)% predicted and PaCO2 52 (4) mm Hg received 6 weeks of nocturnal (1) air (placebo), (2) oxygen and (3) NIV. The primary outcome measures were CF specific QoL, daytime sleepiness and exertional dyspnoea. Secondary outcome measures were awake and asleep gas exchange, sleep architecture, lung function and peak exercise capacity. RESULTS: Compared with air, NIV improved the chest symptom score in the CF QoL Questionnaire (mean difference 10; 95% CI 5 to 16; p = 0.002) and the transitional dyspnoea index score (mean difference 3.1; 95% CI 1.2-5.0; p = 0.01). It reduced maximum nocturnal pressure of transcutaneous CO2 (PtcCO2 mean difference -17 mm Hg; 95% CI -7 to -28 mm Hg; p = 0.005) and increased exercise performance on the Modified Shuttle Test (mean difference 83 m; 95% CI 21 to 144 m; p = 0.02). NIV did not improve sleep architecture, lung function or awake PaCO2. CONCLUSION: 6 weeks of nocturnal NIV improves chest symptoms, exertional dyspnoea, nocturnal hypoventilation and peak exercise capacity in adult patients with stable CF with awake hypercapnia. Further studies are required to determine whether or not NIV can improve survival.

Cytomegalovirus viral load monitoring after allogeneic bone marrow transplantation in patients receiving antiviral prophylaxis.

Cytomegalovirus viral load measurement is a powerful new tool for monitoring of CMV disease; however, the optimal strategy for use is unknown. Weekly plasma CMV viral loads and CMV-related outcomes were monitored in 46 consecutive allogeneic bone marrow transplantation (BMT) recipients receiving standardised antiviral prophylaxis. A total of 412 CMV viral loads were quantitated in the first 100 days post transplantation with 77 positive samples (19%) in 20 patients (43%). No patient with all negative CMV viral load results developed CMV disease. Two of three patients with highly positive CMV viral loads (first positive < or =30 days post transplant, maximum viral load > or =5000 copies/ml, and > or =50% of samples positive) developed CMV disease. A total of 17 patients with positive CMV viral loads, who did not meet the criteria for highly positive, did not develop CMV disease. CMV viral load detection was higher in recipients who were CMV sero-positive. In conclusion, CMV disease did not occur in the setting of a persistently negative CMV viral load. A positive CMV viral load result occurred commonly after allogeneic BMT, even in patients receiving antiviral prophylaxis.

Increased human cytomegalovirus (HCMV) DNA load in peripheral blood leukocytes after lung transplantation correlates with HCMV pneumonitis.

BACKGROUND: Human cytomegalovirus (HCMV) reactivation and disease remain relatively common in lung transplant recipients (LTR) despite the use of ganciclovir prophylaxis protocols for all HCMV at-risk patients. The specific aims of this study were to (1) describe the HCMV DNA viral load in the peripheral blood leukocytes (PBL) of a cohort of LTR during the first 6 months after lung transplantation; (2) prospectively determine whether HCMV DNA viral load predicts episodes of HCMV pneumonitis in LTR; and (3) study the effect of ganciclovir on HCMV viral load. METHODS: Competitive polymerase chain reaction using an internal standard and fluorometric detection were used to quantitate HCMV DNA in the PBL of a cohort of 26 LTR monthly for the first 6 months after transplantation (145 samples). All patients were treated with standard triple immunosuppression, and ganciclovir prophylaxis was given to all at-risk LTR (donor or recipient HCMV seropositive) for at least 8 weeks after transplantation. RESULTS: Thirteen episodes of histopathologically proven HCMV pneumonitis in nine subjects occurred during follow-up with a wide intra- and intersubject variation in the HCMV DNA PBL levels. HCMV detection had a sensitivity of 92% and specificity of 76% for HCMV pneumonitis (negative likelihood ratio, 9.5), whereas greater than 10-fold increases in HCMV DNA load had a specificity of 93% and sensitivity of 67% (positive likelihood ratio, 11). HCMV DNA detection had an adjusted odds ratio for HCMV pneumonitis of 107 (95% confidence interval, 14-821; P<0.005). In those with detectable HCMV DNA in PBL (n=44), HCMV DNA levels were 4.4 (95% confidence interval, 1.2-16.8) times higher in those with HCMV pneumonitis than in those without HCMV pneumonitis. Although ganciclovir treatment was very effective in treating HCMV pneumonitis and suppressing HCMV DNA levels, thrice weekly ganciclovir prophylaxis only partially controlled HCMV DNA levels and did not eliminate HCMV pneumonitis risk as three patients developed HCMV pneumonitis while on this regimen. CONCLUSIONS: HCMV DNA detection, absolute levels, and relative change from baseline in the PBL of LTR correlate with HCMV pneumonitis episodes and may be a useful intermediate outcome measure of the efficacy of ganciclovir prophylaxis and treatment strategies.

Idiopathic pulmonary fibrosis: in need of focused and systematic management.

Idiopathic pulmonary fibrosis (IPF) is an increasingly recognised, serious lung disease. A recent International Consensus Statement has redefined the term "idiopathic pulmonary fibrosis", restricting its use to the entity previously described as "usual interstitial pneumonia" and reclassifying some of the more benign inerstitial lung diseases formerly included under IPF. There is insufficient quality evidence for the effectiveness of current medical therapies for IPF. Lung transplantation provides a potential surgical therapeutic option for selected individuals with IPF, but referral for transplant needs to be made as early as possible. Multidisciplinary clinics specialising in interstitial lung disease have a potential role in determining which patients may benefit from novel and existing medical therapies and which patients should be referred for lung transplantation.

Post-lung transplant bronchiolitis obliterans syndrome (BOS) is characterized by increased exhaled nitric oxide levels and epithelial inducible nitric oxide synthase.

In conditions characterized by airway inflammation, exhaled nitric oxide (eNO) levels are increased. Post-lung transplant bronchiolitis obliterans syndrome (BOS) is characterized by airway inflammation and development of progressive airway narrowing and fibrosis. We have previously shown that in stable lung transplant recipients (LTR), mean eNO levels were not elevated but were still related to the degree of airway neutrophilia within the group. The hypothesis now tested is that in BOS, eNO levels are increased in association with even greater airway neutrophilia and enhanced expression of inducible (iNOS) nitric oxide synthase in the bronchial epithelium. We determined eNO levels in 40 LTR in four groups: well and "stable": LTR (n = 20), BOS (n = 8), bacterial airway infection (BI, n = 6), and acute rejection (AR, n = 6). Following bronchoscopic sampling, we performed a quantitative assessment of iNOS and constitutive nitric oxide synthase (cNOS) expression in endobronchial biopsies by immunohistochemistry. Mean +/- SEM eNO levels in BOS and BI were significantly higher than in stable LTR (20 +/- 1.2 parts per billion [ppb] and 24.7 +/- 1.7 ppb versus 12.5 +/- 0.9 ppb; p < 0.01 for both). In AR, eNO levels (13.4 ppb +/- 0.5) were not different in stable LTR (p = 0.34). When compared with stable LTR, there was increased expression of iNOS in the bronchial epithelium and generally in the lamina propria (LP) in patients with BOS and BI. In AR, iNOS expression was increased but only in the LP in a perivascular distribution. Expression of cNOS was reduced in BOS but not in BI and AR compared with the stable group. Using regression analysis, only iNOS expression in the bronchial epithelium (r(2) = 0.77; p < 0.0001) and %BAL neutrophils (r(2) = 0. 79; p < 0.0001) were positively related to eNO in stable LTR and BOS. We conclude that epithelial iNOS appears to be the major source of eNO. Exhaled NO levels also appear to reflect the degree of airway neutrophilia in both stable LTR and BOS groups. This suggests that serial eNO measurements may be able to predict the early development of BOS.

Pharmacokinetic assessment of oral ganciclovir in lung transplant recipients with cystic fibrosis.

Oral ganciclovir has been used as prophylaxis and therapy against cytomegalovirus in patients with HIV infection and following organ transplantation. Oral ganciclovir has clear practical advantages over intravenous ganciclovir but has a relatively low bioavailability and this may be problematic in at-risk patients with malabsorption. The bioavailability and therefore therapeutic potential of oral ganciclovir in cystic fibrosis (CF) patients post-lung transplant (LT) might be expected to be inadequate given the high incidence of malabsorption in these patients. An 8 h pharmacokinetic study was performed in 12 CF patients 160 +/- 122 days post-transplant who had been taking 1 g oral ganciclovir tds for 3 days with food (plus normal enzyme supplements). Mean (range) serum creatinine was 150 Imol/L (70-280). Blood was sampled at 0.5, 1, 2, 3, 4, 6 and 8 h post-final dose. Plasma was stored at -20 degrees C and later analysed by highperformance liquid chromatography. Mean peak concentration (C(max)) was 4.8 mg/L (0. 96-12.8), mean minimum concentration (C(min)) was 3.6 mg/L (0.78-11. 7) and mean area under the curve (AUC) was 35.4 mg.8 h/L (8-99). C(max), C(min) and AUC correlated significantly with one another (P < 0.001) as well as with serum creatinine and creatinine clearance (P < 0.01). When corrected for alterations in renal function, plasma oral ganciclovir levels are as predicted for other transplant populations. Three days of oral ganciclovir results in therapeutically useful plasma drug levels in the CF LT population, despite a background of general malabsorption. C(max), C(min) and AUC are highly correlated, allowing for the possibility of steady-state drug monitoring to confirm that the recommended dosing algorithm produces appropriate plasma levels.

Bilateral sequential single lung transplantation for pulmonary hypertension and Eisenmenger's syndrome.

BACKGROUND: Lung transplantation, with and without intracardiac repair for pulmonary hypertension (PH) and Eisenmenger's syndrome (EIS), has become an alternative transplant strategy to combined heart and lung transplantation (HLT). METHODS: Thirty-five patients with PH or EIS underwent either bilateral sequential single lung transplantation (BSSLT, group I, n = 13) or HLT (group II, n = 22). Another 74 patients, who underwent BSSLT for other indications, served as controls (group III). Immediate allograft function, early and medium-term outcomes, lung function, and 2-year survival were compared between the groups. RESULTS: Comparisons between groups I and II showed no significant difference in any variables except percent predicted forced vital capacity. Immediate allograft function was significantly inferior (p < 0.05) and the blood loss was greater (p < 0.01) in group I when compared with those in group III. However, this resulted in no significant difference in early and medium-term outcomes, and 2-year survival between the 2 groups. CONCLUSIONS: BSSLT for PH and EIS can be performed as an alternative procedure to HLT without an increase in early and medium-term morbidity and mortality. Results are comparable with BSSLT performed for other indications.

In stable lung transplant recipients, exhaled nitric oxide levels positively correlate with airway neutrophilia and bronchial epithelial iNOS.

In conditions characterized by airway inflammation, exhaled nitric oxide (eNO) levels are increased. Variable degrees of airway inflammation are present in stable lung transplant recipients (LTR), and may lead to airway remodeling and chronic graft dysfunction. The hypothesis tested is that in stable LTR, eNO concentrations would reflect the expression of inducible (iNOS) (but not constitutive [cNOS] nitric oxide synthase) in the bronchial epithelium as well as the degree of airway inflammation. We determined eNO concentrations in 20 stable LTR, free of infection, rejection, or obliterative bronchiolitis (OB). At routine bronchoscopy, we measured the differential cell count on bronchoalveolar lavage (BAL) and a quantitative assessment of iNOS and cNOS expression in endobronchial biopsies by immunohistochemistry. Mean +/- SEM eNO concentrations in stable LTR were not significantly different from control subjects (13 +/- 0.7 ppb versus 14.2 +/- 0.49; p = 0.42). Percent BAL neutrophils was 11.5 +/- 3.2 which was significantly higher than in a group of local control subjects (1.7 +/- 0.6; p < 0.001). The bronchial epithelium and lamina propria contained abundant iNOS but cNOS was present only in the lamina propria. Using regression analysis, percent BAL neutrophils (r(2) = 0.82; p < 0.0001) and iNOS expression in the bronchial epithelium (r(2) = 0.75; p < 0.0001), but not in the lamina propria (r(2) = 0.16; p = 0.08), were positively predictive of eNO. There was an inverse relationship between cNOS and eNO. We conclude that eNO concentrations although normal for the group, still reflect the degree of airway inflammation in stable LTR. Epithelial iNOS appears to be the major source of eNO and expression of cNOS may be downregulated with increasing iNOS expression.

Maximizing the utilization of donor organs offered for lung transplantation.

The number of patients awaiting lung transplantation (LT) and waiting time for surgery is increasing. In Australia, LT rates are 4. 6/million population/yr, which despite low organ donation rates, are the highest published in the world. The Australian organ allocation system allows identification of marginal donors and therapeutic manipulation where appropriate. This study aims to assess the impact of utilization of marginal donors and aggressive donor management. A comparison between published donor criteria and local practice is made, allowing assessment of the effect of using marginal donors on outcome. Donor management included antibiotic therapy, strict fluid management, physiotherapy, bronchoscopy and bronchial toilet, and alteration of ventilatory settings including initiation of pressure support. Blood gases were repeated to assess the results of interventions. Between January 1, 1995 and May 31, 1998, we performed 140 transplants from 112 of 219 (51%) lung donor offers. Of these donors, 48 (43%) satisfied all published criteria for suitable donor organs (Group 1 = ideal donors) and 64 (57%) did not (Group 2 = marginal donors). Criteria breached by the marginal donors were: an initial ratio of arterial oxygen pressure to fraction of inspired oxygen (PaO2/FIO2) < 300 mm Hg (n = 20), abnormal radiology (n = 39), pulmonary infection (n = 24), 20 pack-years smoking (n = 5) and age > 55 yr (n = 4). Therapeutic manipulation resulted in improvement in the PaO2/FIO2 ratio in 20 donors (Group 3) who would not otherwise have been used. Immediate and 24 h postoperative gas exchange and length of intensive care unit (ICU) stay was not different for recipients from donors from all three groups. Overall survival was 94% at 30 d, 83% at 1 yr, 70% at 2 yr, and 62% at 3 yr and was not significantly different from the three groups. We conclude that organ utilization can be maximized by therapeutic manipulation and utilization of marginal donors without compromising results from transplantation.

Impact of graft ischemic time on outcomes after bilateral sequential single-lung transplantation.

BACKGROUND: Graft ischemic time (GIT) is a potential limiting factor in lung transplantation. METHODS: Seventy-four patients who underwent bilateral sequential single-lung transplantation were divided into three groups: group I, GIT less than 5 hours (n = 20); group II, GIT between 5 and 8 hours (n = 39); and group III, GIT more than 8 hours (n = 15). We compared early allograft function (ratio of arterial oxygen tension to inspired oxygen fraction and alveolar-arterial oxygen gradient), blood loss, the need for tracheostomy, the duration of ventilation, intensive care unit stay, and hospital stay. We also compared prevalences of acute and chronic rejection, airway complications, lung function test, and 2-year survival. RESULTS: Early allograft function in group III was significantly worse than those in groups I and II. However, there was no significant difference in any other variables of early and medium-term outcomes among the three groups. No significant correlation was detected between GIT and duration of intensive care unit stay or hospital stay. CONCLUSIONS: The limitation of acceptable GIT could be extended from the traditionally approved 4 to 5 hours, to 5 to 8 hours or even longer.

Expression of the IL-4 receptor alpha-subunit is increased in bronchial biopsy specimens from atopic and nonatopic asthmatic subjects.

BACKGROUND: Recent studies have provided evidence for increased IL-4 expression in the airways of atopic and nonatopic asthmatic subjects. IL-4 is believed to perform important regulatory roles in asthma; however, the expression of the IL-4 receptor has not been investigated. In this study we examined the mRNA and protein expression of the specific alpha-subunit of the IL-4 receptor (alphaIL-4R) in bronchial biopsy specimens obtained from atopic and nonatopic asthmatic subjects. METHODS: Asthmatic subjects and nonasthmatic control subjects were recruited, and lung function measurements were performed before bronchoscopy. Endobronchial biopsy specimens were examined for the presence of alphaIL-4R mRNA and immunoreactivity by using in situ hybridization and immunocytochemistry, respectively. RESULTS: alphaIL-4R mRNA-positive and immunoreactive cells were detected in the epithelium and subepithelium in biopsy specimens from all subjects. Expression of alphaIL-4R mRNA and protein was significantly increased in the epithelium and subepithelium of biopsy specimens from atopic asthmatic subjects compared with atopic control subjects (P <.05 and P <.001, respectively). Epithelial alphaIL-4R mRNA expression and immunoreactivity did not differ significantly between nonatopic asthmatic subjects and nonatopic control subjects. Although the numbers of alphaIL-4R mRNA-positive cells were augmented in the submucosa of intrinsic asthmatic subjects compared with nonatopic control subjects (P <.05), alphaIL-4R immunoreactivity did not differ significantly between these groups. Increased alphaIL-4R immunoreactive signals were also detected in the endothelial cell layer in both atopic and intrinsic asthmatic subjects compared with atopic and nonatopic control subjects, respectively (P <.05). Combined in situ hybridization immunocytochemistry performed on biopsy sections from asthmatic and control subjects demonstrated alphaIL-4R mRNA expression in CD3-positive T cells and tryptasepositive mast cells, with T cells comprising the larger proportion of alphaIL-4R mRNA-positive cells. Numbers of alphaIL-4R mRNA-positive or immunoreactive cells did not correlate with CD3-positive cell numbers, numbers of IL-4 mRNA-positive cells, or indices of pulmonary function. CONCLUSION: These results demonstrate constitutive alphaIL-4R expression in normal airways and enhanced expression in airway tissue from asthmatic individuals.

Inflammation of small airways in asthma.

This study was designed to examine the inflammatory process in the central and peripheral airways of surgically resected lungs from asthmatic and nonasthmatic subjects. Lung specimens were inflated with cryoprotective, rapidly frozen, and systematically sampled. Cryosections prepared from frozen tissue blocks were fixed in acetone/methanol and immunostained with monoclonal antibodies by using the alkaline phosphatase-anti-alkaline phosphatase technique to detect CD3 (T cells), major basic protein (total eosinophils), EG2 (activated eosinophils), anti-tryptase (mast cells), anti-elastase (neutrophils), and CD68 (macrophages). All airways from patients with asthma demonstrated a significant increase in the numbers of T cells and total and activated eosinophils compared with airways from nonasthmatic subjects (p < 0.001). In the patients with asthma, the numbers of activated eosinophils but not T cells were significantly greater in airways with an internal perimeter less than 2 mm compared with those with an internal perimeter greater than 2 mm (p < 0.05). There were also significantly higher numbers of major basic protein-positive eosinophils, when expressed as a fraction of the alveolar wall tissue, in patients with asthma compared with control subjects (p < 0.05). In asthmatic airways with an internal perimeter of more than 2 mm, there was a greater number of activated eosinophils in the tissue between the epithelium and the smooth muscle compared with the tissue between the smooth muscle layer and lung parenchyma (p < 0.05). In contrast, there was a greater number of total eosinophils in the outer airway layer compared with the inner airway layer (p < 0.05). These results show that there is a similar but more severe inflammatory process present in the peripheral compared with the central airways of patients with asthma, which is consistent with the fact that the smaller airways are a major site of obstruction in asthma.

Membrane-bound and soluble alpha IL-5 receptor mRNA in the bronchial mucosa of atopic and nonatopic asthmatics.

Eosinophilic inflammation and interleukin-5 (IL-5) expression are characteristic features of the bronchial mucosa in asthma. We have investigated the differential expression of membrane and soluble isoforms of alpha IL-5 receptor (alpha IL-5Rm and alpha IL-5Rs) mRNA in asthmatics and in normal control subjects and examined the correlation between alpha IL-5Rm and alpha IL-5Rs expression and the FEV1 and airway hyperresponsiveness. Nineteen subjects with stable asthma (atopic = 9; intrinsic = 10) and 22 control subjects (atopic = 12; nonatopic = 10) were recruited. Endobronchial biopsies were obtained and processed for in situ hybridization and double-staining techniques. There was a significant increase in the number of cells per millimeter basement membrane expressing mRNA for total, membrane-bound, and soluble alpha IL-5R in asthmatics when compared with that in nonasthmatic control subjects (p < 0.001); 93% of the cells positive for alpha IL-5R mRNA were EG2+ve eosinophils. There was no significant difference in the expression of alpha IL-5Rm and alpha IL-5Rs between the atopic and nonatopic asthmatics. The expression of alpha IL-5Rm and alpha IL-5Rs was also nonsignificantly different between the atopic and nonatopic control subjects. However, in the asthmatic subjects, the number of positive cells expressing mRNA for alpha IL-5Rm inversely correlated with FEV1(r2 = 0.89, p < 0.001), whereas the expression of alpha IL-5Rs mRNA directly correlated with FEV1 (r2 = 0.52, p < 0.001). There were no significant correlations between alpha IL-5R isoforms and the methacholine PC20. These results suggest that alpha IL-5R upregulation and differential regulation of alternatively spliced alpha IL-5R mRNA transcripts may influence the eosinophil response and the accompanying changes in airflow limitation in both atopic and nonatopic variants of chronic asthma.

IFN-gamma and IL-12 are increased in active compared with inactive tuberculosis.

Cytokine-mediated immune responses to Mycobacterium tuberculosis infection are important determinants of M. tuberculosis disease development and pathology. However, the distinction between changes in cytokine profile attributable to M. tuberculosis infection and those associated with active pulmonary tuberculosis is unclear. We have compared T cells and their subsets, macrophages, and cytokine messenger RNA (mRNA) profile in the bronchoalveolar lavage (BAL) of patients with active pulmonary tuberculosis with inactive tuberculosis subjects. Ten patients with microbiologically confirmed active pulmonary tuberculosis and 25 subjects with inactive tuberculosis were recruited. Bronchoscopy with BAL was undertaken in all cases and BAL cytospins were examined using the techniques of immunocytochemistry and in situ hybridization. There was a significant increase in the percentage of BAL cells that were CD8+ T cells in active tuberculosis compared with inactive tuberculosis (mean +/- SEM: 7.2 +/- 0.9 versus 2.1 +/- 0.4, p < 0.001), but not CD3+ or CD4+ T cells nor macrophages. There were significant increases in the percentage of BAL cells expressing mRNA for interferon-gamma (IFN-gamma) and interleukin-12 (IL-12) in active versus inactive pulmonary tuberculosis subjects (8.0 +/- 0.6 versus 3.7 +/- 0.4 and 28.4 +/- 2.3 versus 10.2 +/- 1.0, p < 0.001, respectively). There were no significant differences between the active and inactive groups in the number of cells expressing mRNA for IL-2, tumor necrosis factor-alpha (TNF-alpha), IL-4, and IL-5. In conclusion, active pulmonary tuberculosis is associated with increased numbers of CD8+ cells and marked increases in the expression of IL-12 and IFN-gamma mRNA in the BAL, both of which may be useful markers of disease activity.

Adoptively transferred late allergic airway responses are associated with Th2-type cytokines in the rat.

Late allergic airway responses can be transferred by CD4+ T cells in the rat. To investigate the role of T-cell cytokines in these responses, we examined the expression of mRNA for Th2 (interleukin [IL]-4 and IL-5) and Th1 (IL-2 and interferon gamma [INF-gamma])-type cytokines in Brown Norway rats that were administered either antigen-primed W3/25(CD4)+ or OX8(CD8)+ T cells. Donors were actively sensitized by subcutaneous injection of ovalbumin (OVA) in the neck and T cells were obtained from the cervical lymph nodes by immunomagnetic cell sorting for administration to unsensitized rats. Control rats received bovine serum albumin (BSA)-primed CD4+ and CD8+ T cells. Two days later, recipient rats were challenged with aerosolized OVA, and bronchoalveolar lavage (BAL) was performed 8 h after challenge. BAL cells expressing mRNA for IL-2, IL-4, IL-5, and INF-gamma were analyzed using the technique of in situ hybridization. Recipients of OVA-primed CD4+ T cells had an increase in the fraction of BAL cells expressing mRNA for IL-4 and IL-5 compared with BSA-primed CD4+ or OVA-primed CD8+ cells (P < 0.001). Recipients of CD8+ T cells had an increase in INF-gamma mRNA expression after OVA challenge compared with recipients of BSA-primed-CD8+ or OVA-primed CD4+ T cells (P < 0.001). In conclusion, T-cell-dependent allergen-induced late responses are associated with the expression of mRNA for IL-4 and IL-5, indicating Th2 cell activation. Furthermore, the increased expression of INF-gamma in allergen challenge recipients of antigen-primed CD8+ T cells suggests that CD8+ T cells may be important in modulating allergic responses.