Multimodal probing of T-cell recognition with hexapod heterostructures.

Studies using antigen-presenting systems at the single-cell and ensemble levels can provide complementary insights into T-cell signaling and activation. Although crucial for advancing basic immunology and immunotherapy, there is a notable absence of synthetic material toolkits that examine T cells at both levels, and especially those capable of single-molecule-level manipulation. Here we devise a biomimetic antigen-presenting system (bAPS) for single-cell stimulation and ensemble modulation of T-cell recognition. Our bAPS uses hexapod heterostructures composed of a submicrometer cubic hematite core (α-Fe2O3) and nanostructured silica branches with diverse surface modifications. At single-molecule resolution, we show T-cell activation by a single agonist peptide-loaded major histocompatibility complex; distinct T-cell receptor (TCR) responses to structurally similar peptides that differ by only one amino acid; and the superior antigen recognition sensitivity of TCRs compared with that of chimeric antigen receptors (CARs). We also demonstrate how the magnetic field-induced rotation of hexapods amplifies the immune responses in suspended T and CAR-T cells. In addition, we establish our bAPS as a precise and scalable method for identifying stimulatory antigen-specific TCRs at the single-cell level. Thus, our multimodal bAPS represents a unique biointerface tool for investigating T-cell recognition, signaling and function.

Oculomotor, Vestibular, and Reaction Time Effects of Sports-Related Concussion: Video-Oculography in Assessing Sports-Related Concussion.

OBJECTIVE: The purpose of the study was to test the ability of oculomotor, vestibular, and reaction time (OVRT) metrics to serve as a concussion assessment or diagnostic tool for general clinical use. SETTING AND PARTICIPANTS: Patients with concussion were high school-aged athletes clinically diagnosed in a hospital setting with a sports-related concussion (n = 50). Control subjects were previously recruited male and female high school student athletes from 3 local high schools (n = 170). DESIGN: Video-oculography was used to acquire eye movement metrics during OVRT tasks, combined with other measures. Measures were compared between groups, and a subset was incorporated into linear regression models that could serve as indicators of concussion. MEASURES: The OVRT test battery included multiple metrics of saccades, smooth pursuit tracking, nystagmoid movements, vestibular function, and reaction time latencies. RESULTS: Some OVRT metrics were significantly different between groups. Linear regression models distinguished control subjects from concussion subjects with high accuracy. Metrics included changes in smooth pursuit tracking, increased reaction time and reduced saccade velocity in a complex motor task, and decreased optokinetic nystagmus (OKN) gain. In addition, optokinetic gain was reduced and more variable in subjects assessed 22 or more days after injury. CONCLUSION: These results indicate that OVRT tests can be used as a reliable adjunctive tool in the assessment of concussion and that OKN results appear to be associated with a prolonged expression of concussion symptoms.

Osteoarthritis is associated with increased failure of proximal femoral fracture fixation.

PURPOSE: The purpose of this study was to evaluate whether the presence of hip osteoarthritis at the time of hip fracture increases treatment failure rates when using either a sliding hip screw (SHS) or proximal femoral nail (PFN) for fracture fixation. METHODS: A retrospective study of a consecutive series of 455 women and 148 men (median age, 83.8 years) treated with SHS or PFN was performed. Osteoarthritis was evaluated based on pre-operative radiographs using the Kellgren and Lawrence grading system. Treatment failure, which was defined as non-union, avascular necrosis, backing out of the implant, cut out of the proximal screws, peri-prosthetic fracture, implant breakage, or conversion to hemi- or total hip arthroplasty, was evaluated for a follow-up period of four to seven years. Optimal placement of the implant (tip-apex distance (TAD) and 3-point fixation) and the effects of age, sex, the quality of reduction, implant type, fracture stability, fracture type, and time to failure were considered confounders of the relationship between failure and osteoarthritis (OA). RESULTS: Among the 32 cases (5.3%) of treatment failure, 12 (2%) showed evidence of osteoarthritis. After controlling for age, sex, the quality of reduction, implant type, fracture stability, fracture type, and TAD, osteoarthritis was associated a greater than threefold increase in treatment failure compared with that of patients without pre-operative evidence of osteoarthritis (OR, 3.26; 95% CI, 1.4-7.65; P = 0.006). CONCLUSIONS: After adjusting for potential confounding factors, radiographic evidence of hip osteoarthritis at the time of hip fracture increases the incidence of treatment failure.

Restoring Cystic Fibrosis Transmembrane Conductance Regulator Function Reduces Airway Bacteria and Inflammation in People with Cystic Fibrosis and Chronic Lung Infections.

RATIONALE: Previous work indicates that ivacaftor improves cystic fibrosis transmembrane conductance regulator (CFTR) activity and lung function in people with cystic fibrosis and G551D-CFTR mutations but does not reduce density of bacteria or markers of inflammation in the airway. These findings raise the possibility that infection and inflammation may progress independently of CFTR activity once cystic fibrosis lung disease is established. OBJECTIVES: To better understand the relationship between CFTR activity, airway microbiology and inflammation, and lung function in subjects with cystic fibrosis and chronic airway infections. METHODS: We studied 12 subjects with G551D-CFTR mutations and chronic airway infections before and after ivacaftor. We measured lung function, sputum bacterial content, and inflammation, and obtained chest computed tomography scans. MEASUREMENTS AND MAIN RESULTS: Ivacaftor produced rapid decreases in sputum Pseudomonas aeruginosa density that began within 48 hours and continued in the first year of treatment. However, no subject eradicated their infecting P. aeruginosa strain, and after the first year P. aeruginosa densities rebounded. Sputum total bacterial concentrations also decreased, but less than P. aeruginosa. Sputum inflammatory measures decreased significantly in the first week of treatment and continued to decline over 2 years. Computed tomography scans obtained before and 1 year after ivacaftor treatment revealed that ivacaftor decreased airway mucous plugging. CONCLUSIONS: Ivacaftor caused marked reductions in sputum P. aeruginosa density and airway inflammation and produced modest improvements in radiographic lung disease in subjects with G551D-CFTR mutations. However, P. aeruginosa airway infection persisted. Thus, measures that control infection may be required to realize the full benefits of CFTR-targeting treatments.

Classifying Adverse Events Following Lower Limb Orthopaedic Surgery in Children With Cerebral Palsy: Reliability of the Modified Clavien-Dindo System.

BACKGROUND: The modified Clavien-Dindo (MCD) system is a reliable tool for classifying adverse events (AEs) in hip preservation surgery and has since been utilized in studies involving lower limb surgery for ambulant and nonambulant children with cerebral palsy (CP). However, the profile of AEs recorded in children with CP compared with typically developing children is different, and the reliability of the MCD in CP is unknown. This study aimed to evaluate the interrater and intrarater reliability of the MCD system for classifying AEs following lower limb surgery in children with CP. METHODS: Eighteen raters were invited to participate, including clinicians from surgical, nursing, and physical therapy professions, and individuals with CP. Following a MCD familiarization session, participants rated 40 clinical scenarios on 2 occasions, 2 weeks apart. Fleiss' κ statistics were used to calculate interrater and intrarater reliability. RESULTS: The overall Fleiss' κ value for interrater reliability in the first rating was 0.70 (95% confidence interval, 0.61-0.80), and increased to 0.75 (95% confidence interval, 0.66-0.84) in the second rating. The average Fleiss' κ value for intrarater reliability was 0.78 (range, 0.48 to 1.00). Grading of more severe AEs (MCD III to V) achieved near perfect agreement (κ, 0.87 to 1.00). There was a lower level of agreement for minor AEs (MCD I-II) (κ, 0.53 to 0.55). A κ score of 0 to 0.2 was deemed as poor, 0.21 to 0.4 as fair, 0.41 to 0.6 as good, 0.61 to 0.8 as very good, and 0.81 to 1.0 as almost perfect agreement. CONCLUSIONS: The MCD System demonstrates a very good interrater and intrarater reliability following lower limb surgery in children with CP. The MCD can be used by clinicians from different health care professions with a high level of reliability. The MCD may improve standardization of AE recording with a view to accurate audits and improved clarity in outcome studies for CP. LEVEL OF EVIDENCE: Level II-diagnostic.

Relationship between pulmonary hyperinflation and dyspnoea severity during acute exacerbations of cystic fibrosis.

BACKGROUND AND OBJECTIVE: Acute exacerbations of cystic fibrosis (CF) occur frequently throughout the course of the disease. Dyspnoea is the most common and distressing symptom experienced by patients during these episodes. We tested the hypothesis that pulmonary hyperinflation is an important determinant of dyspnoea severity during acute exacerbations. METHODS: We studied patients during an acute exacerbation of CF. Lung volumes, spirometry and dyspnoea scores were measured at Day 0, Day 7, at the end of treatment (EOT) and 14 days following the EOT. RESULTS: At the start of treatment, mean residual volume (RV)/total lung capacity (TLC) was 54.9%, which decreased significantly with treatment, as did vital capacity (VC), inspiratory capacity (IC) and dyspnoea scores. IC was the only independent predictor of dyspnoea severity. CONCLUSION: Our study demonstrates significant improvements in hyperinflation, spirometry and dyspnoea scores with treatment of acute exacerbations of CF. Hyperinflation, rather than airflow limitation, may contribute towards the increased dyspnoea during exacerbations.

Macrophage migration inhibitory factor enzymatic activity, lung inflammation, and cystic fibrosis.

RATIONALE: Macrophage migration inhibitory factor (MIF) is a proinflammatory mediator with unique tautomerase enzymatic activity; the precise function has not been clearly defined. We previously demonstrated that individual patients with cystic fibrosis (CF) who are genetically predisposed to be high MIF producers develop accelerated end-organ injury. OBJECTIVES: To characterize the effects of the MIF-CATT polymorphism in patients with CF ex vivo. To investigate the role of MIF's tautomerase activity in a murine model of Pseudomonas aeruginosa infection. METHODS: MIF and tumor necrosis factor (TNF)-α protein levels were assessed in plasma or peripheral blood mononuclear cell (PBMC) supernatants by ELISA. A murine pulmonary model of chronic Pseudomonas infection was used in MIF wild-type mice (mif(+/+)) and in tautomerase-null, MIF gene knockin mice (mif (P1G/P1G)). MEASUREMENTS AND MAIN RESULTS: MIF protein was measured in plasma and PBMCs from 5- and 6-CATT patients with CF; LPS-induced TNF-α production from PBMCs was also assessed. The effect of a specific inhibitor of MIF-tautomerase activity, ISO-1, was investigated in PBMCs. In the murine infection model, total weight loss, differential cell counts, bacterial load, and intraacinar airspace/tissue volume were measured. MIF and TNF-α levels were increased in 6-CATT compared with 5-CATT patients with CF. LPS-induced TNF-α production from PBMCs was attenuated in the presence of ISO-1. In a murine model of Pseudomonas infection, significantly less pulmonary inflammation and bacterial load was observed in mif(P1G/P1G) compared with mif(+/+) mice. CONCLUSIONS: MIF-tautomerase activity may provide a novel therapeutic target in patients with chronic inflammatory diseases such as CF, particularly those patients who are genetically predisposed to produce increased levels of this cytokine.

High-resolution CT of nontuberculous mycobacterium infection in adult CF patients: diagnostic accuracy.

OBJECTIVES: To determine the diagnostic accuracy of high-resolution computed tomography (HRCT) for the detection of nontuberculous mycobacterium infection (NTM) in adult cystic fibrosis (CF) patients. METHODS: Twenty-seven CF patients with sputum-culture-proven NTM (NTM+) underwent HRCT. An age, gender and spirometrically matched group of 27 CF patients without NTM (NTM-) was included as controls. Images were randomly and blindly analysed by two readers in consensus and scored using a modified Bhalla scoring system. RESULTS: Significant differences were seen between NTM (+) and NTM (-) patients in the severity of the bronchiectasis subscore [45 % (1.8/4) vs. 35 % (1.4/4), P = 0.029], collapse/consolidation subscore [33 % (1.3/3 vs. 15 % (0.6/3)], tree-in-bud/centrilobular nodules subscore [43 % (1.7/3) vs. 25 % (1.0/3), P = 0.002] and the total CT score [56 % (18.4/33) vs. 46 % (15.2/33), P = 0.002]. Binary logistic regression revealed BMI, peribronchial thickening, collapse/consolidation and tree-in-bud/centrilobular nodules to be predictors of NTM status (R(2 )= 0.43). Receiver-operator curve analysis of the regression model showed an area under the curve of 0.89, P < 0.0001. CONCLUSION: In adults with CF, seven or more bronchopulmonary segments showing tree-in-bud/centrilobular nodules on HRCT is highly suggestive of NTM colonisation. KEY POINTS: Lung function declines rapidly in cystic fibrosis patients with nontuberculous mycobacterium infection. High-resolution computed tomography can help identify nontuberculous mycobacterium in CF patients. Extensive collapse/consolidation and tree-in-bud/centrilobular nodules are predictive of NTM infection. Multiple bronchopulmonary segments showing tree-in-bud/centrilobular nodules strongly suggest nontuberculous mycobacterium infection.

Duration of intravenous antibiotic treatment for acute exacerbations of cystic fibrosis: A systematic review.

BACKGROUND: Acute exacerbations of Cystic Fibrosis (AECF) are associated with significant morbidity. Recommendations are to treat for 2-3 weeks despite limited data. Spirometry is a measure of clinical response yet appears to plateau at 7-10 days. While durations <9 days have been associated with poorer outcomes, a duration of 10 days may be as effective as 14 days, potentially conferring advantages in terms of cost and adverse events. A 2019 Cochrane review by Abbott et al. did not identify any randomised controlled trials (RCT) comparing durations of treatment. Utilising data from non-randomised studies (NRS), we report a systematic review of intravenous antibiotic treatment, exploring changes in FEV1 (Forced Expiratory Volume in 1 second), CRP (C-reactive protein) and peripheral WBC (white blood cell) count in studies with different treatment durations. STUDY DESIGN AND METHODS: Systematic review of published literature following a search of MEDLINE, Embase, CINAHL and the Cochrane Clinical Trials register. Guidelines from the Preferred Reporting items for Systematic reviews and Meta-Analysis (PRISMA) and reporting Meta-analysis of Observational studies (MOOSE) statement were followed. RESULTS: No randomised controlled trials were identified that specifically examined duration of treatment during AECF. This study included all relevant RCTs and also NRS, grouping according to study characteristics, such as length of treatment, location, year, and also characteristics of the patient population. 52 studies, comprising 79 subgroups, and 1,597 patients, were identified. Mean change (95%CI) in ppFEV1 was 10.13 (9.21-11.05). There was no significant difference in change in ppFEV1 for studies treating for 10-12 days; 8.85 (7.47-10.23), vs 13-15 days; 10.68 (9.53-11.82). Similar changes in CRP and WBC were seen irrespective of treatment duration. CONCLUSION: This systematic review provides evidence that shorter durations of treatment may be associated with similar changes in FEV1, CRP and WBC compared with longer durations.

Acute Pulmonary Exacerbation Phenotypes in Patients with Cystic Fibrosis.

Rationale: The etiology of cystic fibrosis (CF) pulmonary exacerbations (PEx) is likely multifactorial with viral, bacterial, and non-infectious pathways contributing. Objectives: To determine whether viral infection status and CRP (C-reactive protein) can classify subphenotypes of PEx that differ in outcomes and biomarker profiles. Methods: Patients were recruited at time of admission for a PEx. Nasal swabs and sputum samples were collected and processed using the respiratory panel of the FilmArray multiplex polymerase chain reaction (PCR). Serum and plasma biomarkers were measured. PEx were classified using serum CRP and viral PCR: "pauci-inflammatory" if CRP < 5 mg/L, "non-viral with systemic inflammation" if CRP ⩾ 5 mg/L and no viral infection detected by PCR and "viral with systemic inflammation" if CRP ⩾ 5 mg/L and viral infection detected by PCR. Results: Discovery cohort (n = 59) subphenotype frequencies were 1) pauci-inflammatory (37%); 2) non-viral with systemic inflammation (41%); and 3) viral with systemic inflammation (22%). Immunoglobulin G, immunoglobulin M, interleukin-10, interleukin-13, serum calprotectin, and CRP levels differed across phenotypes. Reduction from baseline in forced expiratory volume in 1 second as percent predicted (FEV1pp) at onset of exacerbation differed between non-viral with systemic inflammation and viral with systemic inflammation (-6.73 ± 1.78 vs. -13.5 ± 2.32%; P = 0.025). Non-viral with systemic inflammation PEx had a trend toward longer duration of intravenous antibiotics versus pauci-inflammation (18.1 ± 1.17 vs. 14.8 ± 1.19 days, P = 0.057). There were no differences in percent with lung function recovery to <10% of baseline FEV1pp. Similar results were seen in local and external validation cohorts comparing a pauci-inflammatory to viral/non-viral inflammatory exacerbation phenotypes. Conclusions: Subphenotypes of CF PEx exist with differences in biomarker profile, clinical presentation, and outcomes.

Validation and use of a parametric model for projecting cystic fibrosis survivorship beyond observed data: a birth cohort analysis.

BACKGROUND: The current lifetable approach to survival estimation is favoured by CF registries. Recognising the limitation of this approach, we examined the utility of a parametric survival model to project birth cohort survival estimates beyond the follow-up period, where short duration of follow-up meant median survival estimates were indeterminable. METHODS: Parametric models were fitted to observed survivorship data from the US CF Foundation (CFF) Patient Registry 1980-1994 birth cohort. Model-predicted median survival was estimated. The best fitting model was applied to a Cystic Fibrosis Registry of Ireland dataset to allow an evaluation of the model's ability to estimate predicted median survival. This involved a comparison of birth cohort lifetable predicted and observed (Kaplan-Meier) median survival estimates. RESULTS: A Weibull model with main effects of gender and birth cohort was developed using a US CFF dataset (n=13,115) for which median survival was not directly estimable. Birth cohort lifetable predicted median survival for male and female patients born between 1985 and 1994 and surviving their first birthday was 50.9 and 42.4 years respectively. To evaluate the accuracy of a Weibull model in predicting median survival, a model was developed for the 1980-1984 Cystic Fibrosis Registry of Ireland birth cohort (n=243), which had an observed (Kaplan-Meier) median survival of 27.7 years. Model-predicted median survival estimates were calculated using data censored at different follow-up periods. The estimates converged to the true value as length of follow-up increased. CONCLUSIONS: Accurate prognostic information that is clinically critical for care of patients affected by rare, life-limiting disorders can be provided by parametric survival models. Problems associated with short duration of follow-up for recent birth cohorts can be overcome using this approach, providing better opportunities to monitor survival and plan services locally.

Outcome in cystic fibrosis liver disease.

OBJECTIVES: Evidence suggests that cystic fibrosis liver disease (CFLD) does not affect mortality or morbidity in patients with cystic fibrosis (CF). The importance of gender and age in outcome in CF makes selection of an appropriate comparison group central to the interpretation of any differences in mortality and morbidity in patients with CFLD. METHODS: This is a 7-year follow-up of 42 children with CFLD and their age- and sex-matched controls. Participants were reviewed clinically, biochemically, and radiologically at follow-up. RESULTS: Overall, 85% (72 of 84) of the original cohort were included, 36 CFLD participants and 36 CF controls. There was no significant difference in the number of deaths/transplants between groups (7 of 36 (19.4%) CFLD participants, 3 of 36 (8.3%) CF controls). There was a tendency for participants with CFLD to die younger than their respective CF controls. There was no difference in height, weight, body mass index, or pulmonary function between the groups. Nutritional parameters (sum skinfold thickness 31.6 vs. 42.3, P=0.03; mean upper arm fat area 15.08 vs. 10.59, P=0.001; Shwachman score 43.7 vs. 32.1, P=0.001) were worse among CFLD participants than among CF controls. Cystic fibrosis-related diabetes was more common in CFLD participants (11 of 27 (40.7%) vs. 5 of 33 (15.2%), P=0.02). Eight children (22.2%) with evidence of CFLD at baseline had no clinical evidence of liver disease as adults. CONCLUSIONS: Patients with CFLD have a more severe CF phenotype than do CF patients without liver disease. However, a subgroup of children with CFLD will not manifest clinically significant liver disease as adults.

Fixation eye movement abnormalities and stereopsis recovery following strabismus repair.

We evaluated the effects of strabismus repair on fixational eye movements (FEMs) and stereopsis recovery in patients with fusion maldevelopment nystagmus (FMN) and patients without nystagmus. Twenty-one patients with strabismus, twelve with FMN and nine without nystagmus, were tested before and after strabismus repair. Eye-movements were recorded during a gaze-holding task under monocular viewing conditions. Fast (fixational saccades and quick phases of nystagmus) and slow (inter-saccadic drifts and slow phases of nystagmus) FEMs and bivariate contour ellipse area (BCEA) were analyzed in the viewing and non-viewing eye. Strabismus repair improved the angle of strabismus in subjects with and without FMN, however patients without nystagmus were more likely to have improvement in stereoacuity. The fixational saccade amplitudes and intersaccadic drift velocities in both eyes decreased after strabismus repair in subjects without nystagmus. The slow phase velocities were higher in patients with FMN compared to inter-saccadic drifts in patients without nystagmus. There was no change in the BCEA after surgery in either group. In patients without nystagmus, the improvement of the binocular function (stereopsis), as well as decreased fixational saccade amplitude and intersaccadic drift velocity, could be due, at least partially, to central adaptive mechanisms rendered possible by surgical realignment of the eyes. The absence of improvement in patients with FMN post strabismus repair likely suggests the lack of such adaptive mechanisms in patients with early onset infantile strabismus. Assessment of fixation eye movement characteristics can be a useful tool to predict functional improvement post strabismus repair.

Split anterior tibialis tendon transfer to peroneus brevis for spastic equinovarus in children with hemiplegia.

PURPOSE: The aim of this study is to report the safety and eff-cacy of soft-tissue surgery incorporating split transfer of tibi-alis anterior to peroneus brevis (SPLATT-PB) for children with hemiplegic spastic equinovarus. METHODS: This was a retrospective case series of children and adolescents with spastic hemiplegia who had a novel combination of SPLATT-TB, intramuscular tenotomy of tibialis posterior and either spasticity management or gastrocsole-us lengthening as the index surgery. The principal outcome measures were changes in pain and difficulty with shoe wear and radiological parameters obtained from weight-bearing anteroposterior and lateral radiographs of the affected foot before and after surgery. RESULTS: A total of 63 patients with symptomatic spastic equinovarus met the inclusion criteria. Mean age at surgery was 9.8 years (6 to 18) and the mean follow-up was seven years (range 3 to 10 years). Foot pain and problems with shoe wear improved after surgery. Seven radiological criteria showed a clinically and statistically significant improvement at follow-up, the majority being in the normal range. There were 11 surgical adverse events, all classified as Modified Cla-vien-Dindo Grade II. Three patients required further surgery for recurrent equinus, eight patients required further surgery for valgus deformities and four patients required bony surgery for residual varus deformities. CONCLUSION: Soft-tissue surgery for spastic equinovarus was successful in the majority of children with spastic hemiplegia, particularly between ages eight and 12 years, resulting in a plantigrade, flexible foot with minimal pain or limitations in shoe-wear. Children younger than 8 years at index surgery were more prone to overcorrection into valgus. Children older than 12 years had persistent varus deformities requiring bony surgery. LEVEL OF EVIDENCE: Level IV, retrospective case series.

A clinical investigation into the ability of subjects with lung disease to provide breath specimens using the EvidenzerIRL evidential breath analyser in alcohol intoxicant driving in criminal justice evidence.

The EvidenzerIRL instrument has been in use as an evidential breath analyser in the application of drink driving laws in the Republic of Ireland since 2011. The result of the analysis is used as evidence in prosecutions before the Courts in per se offences of driving under the influence of alcohol as distinct from screening results at the roadside. This study aims to assist doctors, lawyers and judges in assessing drivers' failure to provide valid evidential breath specimens. Since the introduction of the EvidenzerIRL, approximately 10% of evidential breath tests annually result in failure or refusal to provide a successful breath specimen, this is an offence under Irish road traffic laws. The presence of lung disease has been given as a reason for the driver failing to provide evidential breath specimens. The aim of this study is to assess the ability of subjects with lung disease to provide breath specimens using the EvidenzerIRL. Pulmonary function tests (PFT) were carried out on volunteers from outpatients of the pulmonary laboratory in St Vincent's University Hospital, Dublin (n = 58) and a control group with no underlying lung disease (n = 19). After the PFTs all volunteers were asked to provide breath specimens using the EvidenzerIRL. Fourteen (24%) out of 58 lung disease volunteers failed to provide a breath specimen, no one from the control group was unsuccessful. Thirteen females and one male volunteer could not successfully provide. Female volunteers were more likely to fail to provide than male volunteers. A significant difference was found between the median age of successful (62.2 years) and unsuccessful (69.2 years) lung disease volunteers. Only one PFT, percentage predicted of Forced Expiratory Volume in 1 second (FEV1), had a significant difference between the mean of successful (86.6%) and unsuccessful (66.5%) lung disease volunteers. A subject with lung disease was more likely to be successful than unsuccessful. Drivers' effort and operators' guidance through the process were found to be crucial parts to a successful outcome.

EDNRA variants associate with smooth muscle mRNA levels, cell proliferation rates, and cystic fibrosis pulmonary disease severity.

Airway inflammation and pulmonary disease are heterogeneous phenotypes in cystic fibrosis (CF) patients, even among patients with the same cystic fibrosis transmembrane conductance regulator (CFTR) genotype. Endothelin, a proinflammatory peptide and smooth muscle agonist, is increased in CF airways, potentially contributing to the pulmonary phenotype. Four cohorts of CF patients were screened for variants in endothelin pathway genes to determine whether any of these variants associated with pulmonary function. An initial cohort of 808 CF patients homozygous for the common CF mutation, DeltaF508, showed significant association for polymorphisms in the endothelin receptor A gene, EDNRA (P = 0.04), but not in the related endothelin genes (EDN1, EDN2, EDN3, or EDNRB) or NOS1, NOS2A, or NOS3. Variants within EDNRA were examined in three additional cohorts of CF patients, 238 patients from Seattle, WA, 303 from Ireland and the U.K., and 228 from Cleveland, OH, for a total of 1,577 CF patients. The three additional groups each demonstrated a significant association between EDNRA 3'-untranslated region (UTR) variant rs5335 and pulmonary function (P = 0.002). At the molecular level, single nucleotide primer extension assays suggest that the effect of the variants is quantitative. EDNRA mRNA levels from cultured primary tracheal smooth muscle cells are greater for the allele that appears to be deleterious to lung function than for the protective allele, suggesting a mechanism by which increased receptor function is harmful to the CF airway. Finally, cell proliferation studies using human airway smooth muscle cells demonstrated that cells homozygous for the deleterious allele proliferate at a faster rate than those homozygous for the protective allele.