The CD8+ T cell content of transbronchial biopsies from patients with a first episode of clinically stable grade A1 cellular rejection is associated with future chronic lung allograft dysfunction.

BACKGROUND: T cells drive acute cellular rejection (ACR) and its progression to chronic lung allograft dysfunction (CLAD) following lung transplantation. International Society for Heart and Lung Transplantation grade A1 ACR without associated allograft dysfunction is often untreated, yet some patients develop progressive graft dysfunction. T-cell composition of A1 ACR lesions may have prognostic value; therefore, protein-level and epigenetic techniques were applied to transbronchial biopsy tissue to determine whether differential T-cell infiltration in recipients experiencing a first episode of stable grade A1 ACR (StA1R) is associated with early CLAD. METHODS: Sixty-two patients experiencing a first episode of StA1R were divided into those experiencing CLAD within 2 years (n = 13) and those remaining CLAD-free for 5 or more years (n = 49). Imaging mass cytometry (IMC) was used to profile the spectrum and distribution of intragraft T cell phenotypes on a subcohort (n = 16; 8 early-CLAD and 8 no early-CLAD). Immunofluorescence was used to quantify CD4+, CD8+, and FOXP3+ cells. Separately, CD3+ cells were fluorescently labeled, micro-dissected, and the degree of Treg-specific demethylated region methylation was determined. RESULTS: PhenoGraph unsupervised clustering on IMC revealed 50 unique immune cell subpopulations. Methylation and immunofluorescence analyses demonstrated no significant differences in Tregs between early-CLAD and no early-CLAD groups. Immunofluorescence revealed that patients who developed CLAD within 2 years of lung transplantation showed greater CD8+ T cell infiltration compared to those who remained CLAD-free for 5 or more years. CONCLUSIONS: In asymptomatic patients with a first episode of A1 rejection, greater CD8+ T cell content may be indicative of worse long-term outlook.

Association between cytomegalovirus viremia and long-term outcomes in lung transplant recipients.

Although cytomegalovirus (CMV) viremia/DNAemia has been associated with reduced survival after lung transplantation, its association with chronic lung allograft dysfunction (CLAD) and its phenotypes is unclear. We hypothesized that, in a modern era of CMV prophylaxis, CMV DNAemia would still remain associated with death, but also represent a risk factor for CLAD and specifically restrictive allograft syndrome (RAS)/mixed phenotype. This was a single-center retrospective cohort study of all consecutive adult, first, bilateral-/single-lung transplants done between 2010-2016, consisting of 668 patients. Risks for death/retransplantation, CLAD, or RAS/mixed, were assessed by adjusted cause-specific Cox proportional-hazards models. CMV viral load (VL) was primarily modeled as a categorical variable: undetectable, detectable to 999, 1000 to 9999, and ≥10 000 IU/mL. In multivariable models, CMV VL was significantly associated with death/retransplantation (≥10 000 IU/mL: HR = 2.65 [1.78-3.94]; P < .01), but was not associated with CLAD, whereas CMV serostatus mismatch was (D+R-: HR = 2.04 [1.30-3.21]; P < .01). CMV VL was not associated with RAS/mixed in univariable analysis. Secondary analyses with a 7-level categorical or 4-level ordinal CMV VL confirmed similar results. In conclusion, CMV DNAemia is a significant risk factor for death/retransplantation, but not for CLAD or RAS/mixed. CMV serostatus mismatch may have an impact on CLAD through a pathway independent of DNAemia.

Referral rates and barriers to lung transplantation based on pulmonary function criteria in interstitial lung diseases: a retrospective cohort study.

BACKGROUND: Interstitial lung diseases (ILD) unresponsive to medical therapy often require lung transplantation (LTx), which prolongs quality of life and survival. Ideal timing for referral for LTx remains challenging, with late referral associated with significant morbidity and mortality. Among other criteria, patients with ILD should be considered for LTx if forced vital capacity (FVC) is less than 80% or diffusion capacity for carbon monoxide (DLCO) is less than 40%. However, data on referral rates are lacking. OBJECTIVES: To evaluate referral rates for LTx based on pulmonary function tests (PFTs) and identify barriers associated with non-referral. DESIGN: A single-center retrospective cohort study. METHODS: The study consisted of ILD patients who performed PFT between 2014 and 2020. Patients with FVC < 80% or a DLCO < 40% were included in the study. Patients with absolute contraindications to LTx were excluded. Referral rates were computed, and a comparison was made between referred and non-referred subjects. RESULTS: Out of 114 ILD patients meeting criteria for referral to LTx, 35 were referred (30.7%), and 7 proceeded to undergo LTx. Median time from PFT to referral for assessment was 255 days [interquartile range (IQR) 35-1077]. Median time from referral to LTx was 89 days (IQR 59-143). Referred patients were younger (p = 0.003), had lower FVC (p < 0.001), DLCO (p < 0.001), and a higher rate of pulmonary hypertension (p = 0.04). Relatively better PFT, and older age, were significantly associated with non-referral of patients. CONCLUSION: There is under-referral of ILD patients who are eligible for LTx, which is associated with severe disease and missed opportunities for LTx. Further research is required to validate these findings.

Lung transplants: addressing referral gaps for lung disease patientsPatients with severe lung diseases that are unresponsive to medical treatments often require lung transplants to enhance their quality of life and survival. Determining the optimal timing for considering a transplant is challenging, as delaying it can lead to complications. Our study aimed to assess how frequently individuals with lung problems, particularly interstitial lung diseases, were referred for lung transplants based on lung function tests. We conducted a retrospective analysis of medical records for patients with lung diseases who underwent lung function tests between 2014 and 2020. We selected patients whose test results indicated impaired lung function, excluding those who were ineligible for lung transplants due to other medical reasons. Subsequently, we examined the number of patients referred for a lung transplant and compared them to those who were not referred. Our findings revealed that out of 114 patients eligible for a lung transplant, only 35 were referred, representing a referral rate of approximately 31%. Among these, only 7 patients actually underwent the transplant procedure. The time elapsed between the lung function test and the referral for a transplant assessment was notably long, averaging around 255 days. Additionally, once referred, patients waited an average of 89 days for the transplant assessment. Referred patients tended to be younger and had more severe lung disease, characterized by lower lung function test results and a higher likelihood of pulmonary hypertension. Conversely, patients who were not referred generally enjoyed better overall health and were older. This discrepancy highlights the missed opportunities for patients to improve their health and quality of life through lung transplantation. Further research is essential to verify the accuracy of these findings, but this study represents a crucial step toward ensuring that individuals with lung diseases receive the appropriate care they require.

Center variability in the prognostic value of a cumulative acute cellular rejection "A-score" for long-term lung transplant outcomes.

The acute rejection score (A-score) in lung transplant recipients, calculated as the average of acute cellular rejection A-grades across transbronchial biopsies, summarizes the cumulative burden of rejection over time. We assessed the association between A-score and transplant outcomes in 2 geographically distinct cohorts. The primary cohort included 772 double lung transplant recipients. The analysis was repeated in 300 patients from an independent comparison cohort. Time-dependent multivariable Cox models were constructed to evaluate the association between A-score and chronic lung allograft dysfunction or graft failure. Landmark analyses were performed with A-score calculated at 6 and 12 months posttransplant. In the primary cohort, no association was found between A-score and graft outcome. However, in the comparison cohort, time-dependent A-score was associated with chronic lung allograft dysfunction both as a time-dependent variable (hazard ratio, 1.51; P < .01) and when calculated at 6 months posttransplant (hazard ratio, 1.355; P = .031). The A-score can be a useful predictor of lung transplant outcomes in some settings but is not generalizable across all centers; its utility as a prognostication tool is therefore limited.

Persistent Severe Acute Respiratory Syndrome Coronavirus 2 Pneumonia in Patients Treated With Anti-CD20 Monoclonal Antibodies.

We report 8 cases of persistent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pneumonia in patients previously treated with anti-CD20 monoclonal antibodies. Polymerase chain reaction of nasopharyngeal swabs for SARS-CoV-2 was negative in most cases; viral cell cultures confirmed that viable SARS-Co-2 virus was present. Four patients were treated with anti-SARS-CoV-2 hyperimmune globulins with rapid resolution of disease.

Timing of Lung Transplant Referral in Patients with Severe COVID-19 Lung Injury Supported by ECMO.

Severe respiratory failure caused by COVID-19 often requires mechanical ventilation, including extracorporeal membrane oxygenation (ECMO). In rare cases, lung transplantation (LTx) may be considered as a last resort. However, uncertainties remain about patient selection and optimal timing for referral and listing. This retrospective study analyzed patients with severe COVID-19 who were supported by veno-venous ECMO and listed for LTx between July 2020 and June 2022. Out of the 20 patients in the study population, four who underwent LTx were excluded. The clinical characteristics of the remaining 16 patients were compared, including nine who recovered and seven who died while awaiting LTx. The median duration from hospitalization to listing was 85.5 days, and the median duration on the waitlist was 25.5 days. Younger age was significantly associated with a higher likelihood of recovery without LTx after a median of 59 days on ECMO, compared to those who died at a median of 99 days. In patients with severe COVID-19-induced lung damage supported by ECMO, referral to LTx should be delayed for 8-10 weeks after ECMO initiation, particularly for younger patients who have a higher probability of spontaneous recovery and may not require LTx.

Predictors and Adverse Outcomes of Acute Kidney Injury in Hospitalized Renal Transplant Recipients.

Data about in-hospital AKI in RTRs is lacking. We conducted a retrospective study of 292 RTRs, with 807 hospital admissions, to reveal predictors and outcomes of AKI during admission. In-hospital AKI developed in 149 patients (51%). AKI in a previous admission was associated with a more than twofold increased risk of AKI in subsequent admissions (OR 2.13, p < 0.001). Other major significant predictors for in-hospital AKI included an infection as the major admission diagnosis (OR 2.93, p = 0.015), a medical history of hypertension (OR 1.91, p = 0.027), minimum systolic blood pressure (OR 0.98, p = 0.002), maximum tacrolimus trough level (OR 1.08, p = 0.005), hemoglobin level (OR 0.9, p = 0.016) and albumin level (OR 0.51, p = 0.025) during admission. Compared to admissions with no AKI, admissions with AKI were associated with longer length of stay (median time of 3.83 vs. 7.01 days, p < 0.001). In-hospital AKI was associated with higher rates of mortality during admission, almost doubled odds for rehospitalization within 90 days from discharge and increased the risk of overall mortality in multivariable mixed effect models. In-hospital AKI is common and is associated with poor short- and long-term outcomes. Strategies to prevent AKI during admission in RTRs should be implemented to reduce re-admission rates and improve patient survival.

Pulmonary epithelial markers in phenotypes of chronic lung allograft dysfunction.

BACKGROUND: Airway epithelial injury is thought to be a key event in the pathogenesis of chronic lung allograft dysfunction (CLAD). We investigated whether markers of epithelial activity and injury in bronchoalveolar lavage fluid (BAL) correlate with CLAD diagnosis and major CLAD phenotypes: bronchiolitis obliterans syndrome (BOS) vs restrictive allograft syndrome (RAS)-related phenotypes (including RAS, mixed phenotype, and all other patients with RAS-like opacities). METHODS: CLAD status and phenotypes were retrospectively determined in a cohort of all consecutive adult, first, bilateral lung transplants performed 2010-2015, with available BAL samples. All patients with RAS-related phenotypes were included and 1:1 matched with BOS patients based on the time from transplant to CLAD-onset. Subjects who were CLAD-free for a minimum of 3 years post-transplant were 1:1 matched to CLAD patients and included as controls. Proteins that maintain the barrier function of the airway epithelial mucosa (club cell secretory protein, surfactant protein-D and epithelial mucins: MUC1, MUC5AC, MUC5B, MUC16), as well as epithelial cell death markers (M30&M65 representing epithelial cell apoptosis and overall death, respectively), were measured in BAL obtained within 6-months post CLAD onset using a double-sandwich ELISA or a multiplex bead assay. Protein levels were compared using Mann-Whitney-U-test. Association between protein levels and graft survival was assessed using Cox proportional hazards models, adjusted for CMV serology mismatch status and CLAD phenotype. RESULTS: Fifty-four CLAD (27 BOS, 11 RAS, 7 mixed, 9 others with RAS-like opacities) patients and 23 CLAD-free controls were included. Median BAL levels were significantly higher in patients with CLAD compared to CLAD-free controls for M30 (124.5 vs 88.7 U/L), MUC1 (6.8 vs 3.2 pg/mL), and MUC16 (121.0 vs 30.1 pg/mL). When comparing CLAD phenotypes, M30 was significantly higher in patients with RAS-related phenotypes than BOS (160.9 vs 114.6 U/L). In multivariable models, higher M30 and MUC5B levels were associated with decreased allograft survival after CLAD onset independent of phenotype (p < 0.05 for all). CONCLUSIONS: Airway epithelial mucins and cell death markers are enhanced in the BAL of patients with CLAD and can assist in differentiating between CLAD phenotypes and post-CLAD outcomes. Abnormal airway mucin expression and epithelial cell death may be involved in the pathogenesis of CLAD, and therefore their detection may aid in future selection of targeted therapies.

Utility of bile acids in large airway bronchial wash versus bronchoalveolar lavage as biomarkers of microaspiration in lung transplant recipients: a retrospective cohort study.

BACKGROUND: Bronchoalveolar lavage (BAL) is a key tool in respiratory medicine for sampling the distal airways. BAL bile acids are putative biomarkers of pulmonary microaspiration, which is associated with poor outcomes after lung transplantation. Compared to BAL, large airway bronchial wash (LABW) samples the tracheobronchial space where bile acids may be measurable at more clinically relevant levels. We assessed whether LABW bile acids, compared to BAL bile acids, are more strongly associated with poor clinical outcomes in lung transplant recipients. METHODS: Concurrently obtained BAL and LABW at 3 months post-transplant from a retrospective cohort of 61 lung transplant recipients were analyzed for taurocholic acid (TCA), glycocholic acid (GCA), and cholic acid by mass spectrometry and 10 inflammatory proteins by multiplex immunoassay. Associations between bile acids with inflammatory proteins and acute lung allograft dysfunction were assessed using Spearman correlation and logistic regression, respectively. Time to chronic lung allograft dysfunction and death were evaluated using multivariable Cox proportional hazards and Kaplan-Meier methods. RESULTS: Most bile acids and inflammatory proteins were higher in LABW than in BAL. LABW bile acids correlated with inflammatory proteins within and between sample type. LABW TCA and GCA were associated with acute lung allograft dysfunction (OR = 1.368; 95%CI = 1.036-1.806; P = 0.027, OR = 1.064; 95%CI = 1.009-1.122; P = 0.022, respectively). No bile acids were associated with chronic lung allograft dysfunction. Adjusted for risk factors, LABW TCA and GCA predicted death (HR = 1.513; 95%CI = 1.014-2.256; P = 0.042, HR = 1.597; 95%CI = 1.078-2.366; P = 0.020, respectively). Patients with LABW TCA in the highest tertile had worse survival compared to all others. CONCLUSIONS: LABW bile acids are more strongly associated than BAL bile acids with inflammation, acute lung allograft dysfunction, and death in lung transplant recipients. Collection of LABW may be useful in the evaluation of microaspiration in lung transplantation and other respiratory diseases.

Physiological Effects of Wearing N95 Respirator on Medical Staff During Prolong Work Hours in Covid-19 Departments.

OBJECTIVE: The objective of the current study was to determine gas exchange abnormalities and physiological changes among healthcare workers during a 4-hour emergency department (ED) shift while wearing the N95 respirator. METHODS: Single-center prospective observational study. Comparisons of paired measurements were performed using a non-parametric Wilcoxon matched-pairs signed-rank test. Results: Forty-one subjects were included. Prolonged N95 respirator use was associated with a significant decline in plasma pH (7.35 mmHg vs 7.34 mmHg, P = 0.02), PvO 2 (23.2 mmHg vs 18.6 mmHg, P < 0.001) and a concurrent increase in EtCO 2 (32.5 mmHg vs 38.5 mmHg, P < 0.0001). PvCO 2 and bicarbonate levels did not differ. No significant change was observed for heart rate or oxygen saturation. CONCLUSION: Using an N95 respirator for prolonged periods by healthcare professionals may provoke changes in gas exchange. The clinical significance of these changes remains to be determined.

Phenotyping CLAD after single lung transplant: Limits and prognostic assessment of the 2019 ISHLT classification system.

BACKGROUND: Phenotyping chronic lung allograft dysfunction (CLAD) in single lung transplant (SLTX) is challenging, due to the native lung contribution to pulmonary function test (PFT). We aimed to assess the applicability and prognostic performance of International Society for Heart and Lung Transplantation (ISHLT) classification in SLTX. METHODS: In this retrospective study of adult, first, SLTX performed 2009-2017, patients with persistent drop in FEV1≥20% were assessed by 2 independent adjudicators to determine CLAD status and phenotype. Interobserver agreement (IOA) was calculated (Cohen's Kappa) for CLAD, phenotype and presence of RAS (resttrictive allograft syndrome)-like opacities (RLO). Association of CLAD phenotypes with time to death or retransplant (ReTx), adjusted for age at SLTX, sex, CMV mismatch and native lung condition, were assessed using Cox proportional hazards models. RESULTS: Of 172 SLTX recipients, 92 experienced a persistent drop in FEV1>20%. Following adjudication, 67 were diagnosed with CLAD. We noted a moderate IOA for CLAD diagnosis (Kappa 0.69) and poor IOA for phenotype adjudication (Kappa 0.52). The final phenotype adjudication was 31 bronchiolitis obliterans syndrome (BOS) (46.3%), 13 RAS (19.4%), 2 mixed (3%), 2 Undefined (3%), and 19 remained Unclassified (28.3%). Using these adjudicated phenotypes, RAS was significantly associated with a higher risk of death/ReTx compared to other groups (HR 2.98, 95%CI [1.39-6.4]). The adjudication of RLO had the best IOA (Kappa 0.73). The presence of RLO was a strong predictor of death or ReTx (HR 2.37, 95%CI [1.2-4.5]), regardless of the final phenotype. CONCLUSIONS: PFT interpretation is challenging in SLTX. A classification essentially relying on imaging, which harbored good IOA, obtained better prognostic performance than a classification using published physiological cut-offs.

Chronic lung allograft dysfunction phenotype and prognosis by machine learning CT analysis.

BACKGROUND: Chronic lung allograft dysfunction (CLAD) is the principal cause of graft failure in lung transplant recipients and prognosis depends on CLAD phenotype. We used a machine learning computed tomography (CT) lung texture analysis tool at CLAD diagnosis for phenotyping and prognostication compared with radiologist scoring. METHODS: This retrospective study included all adult first double lung transplant patients (January 2010-December 2015) with CLAD (censored December 2019) and inspiratory CT near CLAD diagnosis. The machine learning tool quantified ground-glass opacity, reticulation, hyperlucent lung and pulmonary vessel volume (PVV). Two radiologists scored for ground-glass opacity, reticulation, consolidation, pleural effusion, air trapping and bronchiectasis. Receiver operating characteristic curve analysis was used to evaluate the diagnostic performance of machine learning and radiologist for CLAD phenotype. Multivariable Cox proportional hazards regression analysis for allograft survival controlled for age, sex, native lung disease, cytomegalovirus serostatus and CLAD phenotype. RESULTS: 88 patients were included (57 bronchiolitis obliterans syndrome (BOS), 20 restrictive allograft syndrome (RAS)/mixed and 11 unclassified/undefined) with CT a median 9.5 days from CLAD onset. Radiologist and machine learning parameters phenotyped RAS/mixed with PVV as the strongest indicator (area under the curve (AUC) 0.85). Machine learning hyperlucent lung phenotyped BOS using only inspiratory CT (AUC 0.76). Radiologist and machine learning parameters predicted graft failure in the multivariable analysis, best with PVV (hazard ratio 1.23, 95% CI 1.05-1.44; p=0.01). CONCLUSIONS: Machine learning discriminated between CLAD phenotypes on CT. Both radiologist and machine learning scoring were associated with graft failure, independent of CLAD phenotype. PVV, unique to machine learning, was the strongest in phenotyping and prognostication.

Significance of phenotype change after chronic lung allograft dysfunction onset.

Definitions for chronic lung allograft dysfunction (CLAD) phenotypes were recently revised (2019 ISHLT consensus). Post-CLAD onset phenotype transition may occur as a result of change in obstruction, restriction, or RAS-like opacities (RLO). We aimed to assess the prevalence and prognostic implications of these transitions. This was a single-center, retrospective cohort study of bilateral lung transplants performed in 2009-2015. CLAD phenotypes were determined per ISHLT guidelines. CLAD phenotype transition was defined as a sustained change in obstruction, restriction or RLO. We specifically focused on phenotype changes based on RLO emergence. Association of RLO development with time to death or retransplant were assessed using Kaplan-Meier and Cox proportional hazards models. Among 211 patients with CLAD, 47 (22.2%) experienced a phenotype transition. Nineteen patients developed RLO. Development of RLO phenotype after CLAD onset was associated with a shorter time to death/retransplant when considering the entire CLAD patient cohort (HR = 4.00, CI 2.74-5.83, P < 0.001) and also when restricting the analysis to only patients with a Non-RLO phenotype at CLAD onset (HR 9.64, CI 5.52-16.84, P < 0.0001). CLAD phenotype change based on emergence of RAS-like opacities implies a worse outcome. This highlights the clinical importance of imaging follow-up to monitor for phenotype transitions after CLAD onset.

Bronchoalveolar lavage cytokine-based risk stratification of minimal acute rejection in clinically stable lung transplant recipients.

BACKGROUND: Acute cellular rejection (ACR) remains the most significant risk factor for chronic lung allograft dysfunction (CLAD). While clinically significant or higher-grade (≥A2) ACR is generally treated with augmented immunosuppression (IS), the management of clinically stable grade A1 ACR remains controversial. At our center, patients with clinically stable grade A1 ACR are routinely not treated with augmented IS. While the overall outcomes in this group of patients at our center are equivalent to patients with stable A0 pathology, CLAD and death rates remain overall high. We hypothesized that a distinct cytokine signature at the time of early minimal rejection state would be associated with worse outcomes. Specifically, we aimed to determine whether bronchoalveolar lavage (BAL) biomarkers at the time of first clinically stable grade A1 ACR (CSA1R) are predictive of subsequent CLAD or death. METHODS: Among all adult, bilateral, first lung transplants, performed 2010-2016, transbronchial biopsies obtained within the first-year post-transplant were categorized as clinically stable or unstable based on the presence or absence of ≥10% concurrent drop in forced expiratory volume in 1 second (FEV1). We assessed BAL samples obtained at the time of CSA1R episodes, which were not preceded by another ACR (i.e., first episodes). Twenty-one proteins previously associated with ACR or CLAD were measured in the BAL using a multiplex bead assay. Association between protein levels and subsequent CLAD or death was assessed using Cox Proportional Hazards models, adjusted for relevant peri-transplant clinical covariates. RESULTS: We identified 75 patients with first CSA1R occurring at a median time of 98 days (range 48.5-197) post-transplant. Median time from transplant to CLAD or death was 1247 (756.5-1921.5) and 1641 days (1024.5-2326.5), respectively. In multivariable models, levels of MCP1/CCL2, S100A8, IL10, TNF-receptor 1, and pentraxin 3 (PTX3) were associated with both CLAD development and death (p < 0.05 for all). PTX3 remained significantly associated with both CLAD and death after adjusting for multiple comparisons. CONCLUSION: Our data indicate that a focused BAL protein signature, with PTX3 having the strongest association, may be useful in determining a subset of CSA1R patients at increased risk and may benefit from a more aggressive management strategy.

Association between the renin-angiotensin system and chronic lung allograft dysfunction.

Chronic lung allograft dysfunction (CLAD) is the major cause of death after lung transplantation. Angiotensin II (AngII), the main effector of the renin-angiotensin system, elicits fibrosis in both kidney and lung. We identified six AngII-regulated proteins (Ras homolog family member B (RHOB), bone marrow stromal cell antigen 1 (BST1), lysophospholipase 1 (LYPA1), glutamine synthetase (GLNA), thrombospondin 1 (TSP1) and laminin subunit ß2 (LAMB2)) that were increased in urine of patients with kidney allograft fibrosis. We hypothesised that the renin-angiotensin system is active in CLAD and that AngII-regulated proteins are increased in bronchoalveolar lavage fluid (BAL) of CLAD patients.We performed immunostaining of AngII receptors (AGTR1 and AGTR2), TSP1 and GLNA in 10 CLAD lungs and five controls. Using mass spectrometry, we quantified peptides corresponding to AngII-regulated proteins in BAL of 40 lung transplant recipients (stable, acute lung allograft dysfunction (ALAD) and CLAD). Machine learning algorithms were developed to predict CLAD based on BAL peptide concentrations.Immunostaining demonstrated significantly more AGTR1+ cells in CLAD versus control lungs (p=0.02). TSP1 and GLNA immunostaining positively correlated with the degree of lung fibrosis (R2=0.42 and 0.57, respectively). In BAL, we noted a trend towards higher concentrations of AngII-regulated peptides in patients with CLAD at the time of bronchoscopy, and significantly higher concentrations of BST1, GLNA and RHOB peptides in patients that developed CLAD at follow-up (p<0.05). The support vector machine classifier discriminated CLAD from stable and ALAD patients at the time of bronchoscopy (area under the curve (AUC) 0.86) and accurately predicted subsequent CLAD development (AUC 0.97).Proteins involved in the renin-angiotensin system are increased in CLAD lungs and BAL. AngII-regulated peptides measured in BAL may accurately identify patients with CLAD and predict subsequent CLAD development.

IL-6 receptor blockade for allograft dysfunction after lung transplantation in a patient with COPA syndrome.

OBJECTIVE: COPA syndrome is a genetic disorder of retrograde cis-Golgi vesicle transport that leads to upregulation of pro-inflammatory cytokines (mainly IL-1ß and IL-6) and the development of interstitial lung disease (ILD). The impact of COPA syndrome on post-lung transplant (LTx) outcome is unknown but potentially detrimental. In this case report, we describe progressive allograft dysfunction following LTx for COPA-ILD. Following the failure of standard immunosuppressive approaches, detailed cytokine analysis was performed with the intention of personalising therapy. METHODS: Multiplexed cytokine analysis was performed on serum and bronchoalveolar lavage (BAL) fluid obtained pre- and post-LTx. Peripheral blood mononuclear cells (PMBCs) obtained pre- and post-LTx were stimulated with PMA, LPS and anti-CD3/CD28 antibodies. Post-LTx endobronchial biopsies underwent microarray-based gene expression analysis. Results were compared to non-COPA LTx recipients and non-LTx healthy controls. RESULTS: Multiplexed cytokine analysis showed rising type I/II IFNs, and IL-6 in BAL post-LTx that decreased following treatment of acute rejection but rebounded with further clinical deterioration. In vitro stimulation of PMBCs suggested that myeloid cells were driving deterioration, through IL-6 signalling pathways. Tocilizumab (IL-6 receptor antibody) administration for 3 months (4 mg kg-1, monthly) effectively suppressed IL-6 levels in BAL. Mucosal gene expression profile following tocilizumab suggested greater similarity to normal. CONCLUSION: Clinical effectiveness of IL-6 receptor blockade was not observed. However, we identified IL-6 upregulation associated with graft injury, effective IL-6 suppression with tocilizumab and evidence of beneficial effect on molecular transcripts. This mechanistic analysis suggests a role for IL-6 blockade in post-LTx care that should be investigated further.

The Impact of Inadequate ("AX") Transbronchial Biopsies on Post-lung Transplant CLAD or Death.

BACKGROUND: Procuring a good quality transbronchial-biopsy sample is essential for diagnosing acute cellular rejection after lung transplantation (LT). Insufficient transbronchial-biopsy samples are graded "AX." We hypothesized that AX samples may be associated with a higher risk for chronic lung allograft dysfunction (CLAD) or death/retransplant, through a potential anatomic or physiologic underlying pulmonary process or because of undiagnosed acute cellular rejection episodes. METHODS: We conducted a single-center, retrospective, cohort study drawn from all consecutive adult, first, bilateral LT between 1999 and 2015. We reviewed all biopsies obtained within the first year posttransplant and compared outcomes of patients with ≥1 AX to patients with no AX. Association of any AX or percent AX with time to CLAD or death/retransplant was assessed using Cox Proportional Hazards models. RESULTS: The cohort consisted of 809 patients with a median of 6 (interquartile range 5-6) biopsies and 16.7% (interquartile range 0-25) AX samples within the first year posttransplant. Four hundred thirty-nine (54.3%) subjects had ≥1 AX sample obtained within the time period. Median time to CLAD or death/retransplant, from 1 year posttransplant, was 761 (320, 1587) and 1200 (662, 2308) days, respectively. In the multivariable analysis, there was no difference in risk for CLAD (hazard ratio = 1.05, 95% confidence interval, 0.87-1.28, P = 0.60), or death/retransplant (hazard ratio = 1.14, 95% confidence interval, 0.92-1.42, P = 0.24) between patients with ≥1 AX biopsy versus none. Among subjects with ≥1 AX, having >50% AX biopsies was not associated with outcome. CONCLUSIONS: This is the first study to demonstrate that AX biopsies are not associated with an increased risk of CLAD or death/retransplant after LT and may not require to repeat the biopsy.

Eosinophils in transbronchial biopsies: a predictor of chronic lung allograft dysfunction and reduced survival after lung transplantation - a retrospective single-center cohort study.

Long-term outcomes after lung transplantation remain inferior to those of other solid organ groups. The significance of eosinophils detected on transbronchial biopsies (TBBx) after lung transplantation and their relationship to long-term outcomes remain unknown. A retrospective single-center cohort study was performed of patients transplanted between January 01, 2001, and July 31, 2018, who had at least 1 TBBx with evaluable parenchymal tissue. Multivariable Cox proportional hazard models were used to assess the associations between eosinophil detection and: all-cause mortality and Chronic Lung Allograft Dysfunction (CLAD). 8887 TBBx reports from 1440 patients were reviewed for the mention of eosinophils in the pathology report. 112 (7.8%) patients were identified with eosinophils on at least one TBBx. The median (95% CI) survival time for all patients was 8.28 (7.32-9.31) years. Multivariable analysis, adjusted for clinical variables known to affect post-transplant outcomes, showed that the detection of eosinophils was independently associated with an increased risk of death (HR 1.51, 95% CI 1.24-1.85, p < 0.01) and CLAD (HR 1.35, 95% CI 1.07-1.70, P = 0.01). Eosinophils detected in TBBx are associated with an increased risk of CLAD and death. There may be benefit in specifically reporting the presence of eosinophils in TBBx reports and incorporating their presence in clinical decision-making.