Distinct Non-Human Leukocyte Antigen Antibody Signatures Correlate with Endothelial Crossmatch Status in Lung and Renal Transplant Recipients.

Non-HLA antibodies against heterogeneous targets on endothelial cells have been associated with allograft injuries. The endothelial cell crossmatch (ECXM) is used in the detection of non-HLA antibodies but remains non-discriminatory for specific antibody identification. The primary objective of this study was to delineate the specific non-HLA antibody signatures associated with ECXM positivity and to determine the correlation of ECXM status and non-HLA antibody signatures on allograft health. Serum specimens from 25 lung transplant recipients (LTRs) and 13 renal transplant recipients (RTRs) were collected as part of clinical evaluation, and testing for angiotensin II receptor type 1 (AT1R) and donor-specific MHC class I chain-related gene A (MICA) antibodies and ECXM was performed. Remnant sera were tested for non-HLA antibodies using the LABScreen™ Autoantibody (LSAUT) Group 1, 2, and 3 kits (One Lambda, Inc., Los Angeles, CA, USA). In both cohorts, the concordance of AT1R and MICA together or individually with ECXM+ status was poor (<0.7), suggesting the presence of other unaccounted antibodies. Autoantibody profiling revealed three distinct clusters targeting fibrotic products, cytoskeletal proteins, and cell signaling molecules. A comparative analysis of ECXM+ and ECXM- specimens identified nine and five differentially expressed antibodies in the LTR and RTR cohorts, respectively. Employing machine learning techniques (variable importance, feature selection, ROC-AUC), we derived a five-antibody panel (TNFα, collagen V, CXCL11, GDNF, GAPDH) and a two-antibody panel (TNFα, CXCL9) that effectively discriminated between ECXM+ and ECXM- status in the LTR and RTR cohorts, respectively. Distinct antibody signatures were identified in LTR and RTR cohorts that correlated with ECXM+ status and were associated with allograft dysfunction.

Differences in airway microbiome and metabolome of single lung transplant recipients.

BACKGROUND: Recent studies suggest that alterations in lung microbiome are associated with occurrence of chronic lung diseases and transplant rejection. To investigate the host-microbiome interactions, we characterized the airway microbiome and metabolome of the allograft (transplanted lung) and native lung of single lung transplant recipients. METHODS: BAL was collected from the allograft and native lungs of SLTs and healthy controls. 16S rRNA microbiome analysis was performed on BAL bacterial pellets and supernatant used for metabolome, cytokines and acetylated proline-glycine-proline (Ac-PGP) measurement by liquid chromatography-high-resolution mass spectrometry. RESULTS: In our cohort, the allograft airway microbiome was distinct with a significantly higher bacterial burden and relative abundance of genera Acinetobacter & Pseudomonas. Likewise, the expression of the pro-inflammatory cytokine VEGF and the neutrophil chemoattractant matrikine Ac-PGP in the allograft was significantly higher. Airway metabolome distinguished the native lung from the allografts and an increased concentration of sphingosine-like metabolites that negatively correlated with abundance of bacteria from phyla Proteobacteria. CONCLUSIONS: Allograft lungs have a distinct microbiome signature, a higher bacterial biomass and an increased Ac-PGP compared to the native lungs in SLTs compared to the native lungs in SLTs. Airway metabolome distinguishes the allografts from native lungs and is associated with distinct microbial communities, suggesting a functional relationship between the local microbiome and metabolome.

Nanoparticle drug delivery characterization for fluticasone propionate and in vitro testing 1.

Glucocorticoids, such as fluticasone propionate (FP), are used for the treatment of inflammation and alleviation of nasal symptoms and allergies, and as an antipruritic. However, both short- and long-term therapeutic use of glucocorticoids can lead to muscle weakness and atrophy. In the present study, we evaluated the feasibility of the nanodelivery of FP with poly(dl-lactide-co-glycolide) (PLGA) and tested in vitro function. FP-loaded PLGA nanoparticles were prepared via nanoprecipitation and morphological characteristics were studied via scanning electron microscopy. FP-loaded nanoparticles demonstrated an encapsulation efficiency of 68.6% ± 0.5% with a drug loading capacity of 4.6% ± 0.04%, were 128.8 ± 0.6 nm in diameter with a polydispersity index of 0.07 ± 0.008, and displayed a zeta potential of -19.4 ± 0.7. A sustained in vitro drug release pattern was observed for up to 7 days. The use of fluticasone nanoparticle decreased lipopolysaccharide (LPS)-induced lactate dehydrogenase release compared with LPS alone in C2C12 treated cells. FP also decreased expression of LPS-induced inflammatory genes in C2C12 treated cells as compared with LPS alone. Taken together, the present study demonstrates in vitro feasibility of PLGA-FP nanoparticle delivery to the skeletal muscle cells, which may be beneficial for treating inflammation.

The neutrophil chemoattractant peptide proline-glycine-proline is associated with acute respiratory distress syndrome.

Acute respiratory distress syndrome (ARDS) is characterized by unrelenting polymorphonuclear neutrophil (PMN) inflammation and vascular permeability. The matrikine proline-glycine-proline (PGP) and acetylated PGP (Ac-PGP) have been shown to induce PMN inflammation and endothelial permeability in vitro and in vivo. In this study, we investigated the presence and role of airway PGP peptides in acute lung injury (ALI)/ARDS. Pseudomonas aeruginosa-derived lipopolysaccharide (LPS) was instilled intratracheally in mice to induce ALI, and increased Ac-PGP with neutrophil inflammation was noted. The PGP inhibitory peptide, arginine-threonine-arginine (RTR), was administered (it) 30 min before or 6 h after LPS injection. Lung injury was evaluated by detecting neutrophil infiltration and permeability changes in the lung. Pre- and posttreatment with RTR significantly inhibited LPS-induced ALI by attenuating lung neutrophil infiltration, pulmonary permeability, and parenchymal inflammation. To evaluate the role of PGP levels in ARDS, minibronchoalveolar lavage was collected from nine ARDS, four cardiogenic edema, and five nonlung disease ventilated patients. PGP levels were measured and correlated with Acute Physiology and Chronic Health Evaluation (APACHE) score, PaO2 to FIO2 (P/F), and ventilator days. PGP levels in subjects with ARDS were significantly higher than cardiogenic edema and nonlung disease ventilated patients. Preliminary examination in both ARDS and non-ARDS populations demonstrated PGP levels significantly correlated with P/F ratio, APACHE score, and duration on ventilator. These results demonstrate an increased burden of PGP peptides in ARDS and suggest the need for future studies in ARDS cohorts to examine correlation with key clinical parameters.

Leukemia inhibitory factor (LIF) receptor amplifies pathogenic activation of fibroblasts in lung fibrosis.

Fibrosis drives end-organ damage in many diseases. However, clinical trials targeting individual upstream activators of fibroblasts, such as TGFß, have largely failed. Here, we target the leukemia inhibitory factor receptor (LIFR) as a "master amplifier" of multiple upstream activators of lung fibroblasts. In idiopathic pulmonary fibrosis (IPF), the most common fibrotic lung disease, we found that lung myofibroblasts had high LIF expression. Further, TGFß1, one of the key drivers of fibrosis, upregulated LIF expression in IPF fibroblasts. In vitro anti-LIFR antibody blocking on human IPF lung fibroblasts reduced induction of profibrotic genes downstream of TGFß1, IL-4 and IL-13. Further, siRNA silencing of LIFR in IPF precision cut lung slices reduced expression of fibrotic proteins. Together, we find that LIFR drives an autocrine positive feedback loop that amplifies and sustains pathogenic activation of IPF fibroblasts downstream of multiple external stimuli, implicating LIFR as a therapeutic target in fibrosis. Significance Statement: Fibroblasts have a central role in the pathogenesis of fibrotic diseases. However, due to in part to multiple profibrotic stimuli, targeting a single activator of fibroblasts, like TGFß, has not yielded successful clinical treatments. We hypothesized that a more effective therapeutic strategy is identifying a downstream "master amplifier" of a range of upstream profibrotic stimuli. This study identifies the leukemia inhibitory factor receptor (LIFR) on fibrotic lung fibroblasts amplifies multiple profibrotic stimuli, such as IL-13 and TGFß. Blocking LIFR reduced fibrosis in ex vivo lung tissue from patients with idiopathic pulmonary fibrosis (IPF). LIFR, acting as a master amplifier downstream of fibroblast activation, offers an alternative therapeutic strategy for fibrotic diseases.

Esophageal Function and Reflux Evaluations in Lung Transplantation: A Nationwide Survey of UNOS-Accredited Transplant Centers in the United States.

INTRODUCTION: Gastroesophageal reflux disease has been associated with worse lung transplant outcomes. We aimed to assess local practices for esophageal function testing (EFT) across transplant centers. METHODS: This was a survey study of all United Network for Organ Sharing-accredited adult lung transplant centers regarding local EFT practice. RESULTS: Among 39/63 (60%) responded centers, 38.5% required any EFT (35.9% esophageal manometry, 15.4% pH monitoring, and 28.2% pH impedance), while another 28.2% may consider EFT based on symptoms. Five-year transplant volume was higher among centers requiring EFT (253 vs 159, P = 0.04). DISCUSSION: Only a minority of lung transplant centers routinely obtained EFT, supporting the need for guidelines for standardized reflux/esophageal assessment.

Adverse Outcomes Associated With Atrial Arrhythmias After Veno-Venous Extracorporeal Membrane Oxygenation.

Veno-venous extracorporeal membrane oxygenation (VV ECMO) is used as a treatment modality in those who fail to respond to conventional care. Hypoxia and medications used in the intensive care unit may increase risk for atrial arrhythmias (AA). This study aims to evaluate the impact of AA on post-VV ECMO outcome. A retrospective review of patients who were placed on VV ECMO between October 2016 and October 2021. One hundred forty-five patients were divided into two groups, AA and no AA. Baseline characteristic and potential risk factors were assessed. Uni- and multivariate analysis using logistic regression models were constructed to evaluate the predictors of mortality between groups. Survival between groups was estimated by the Kaplan-Meier method using the log-rank test. Advanced age with history of coronary artery disease and hypertension were associated with increased risk to develop AA post-VV ECMO placement ( p value < 0.05). Length on ECMO, time intubated, hospital length of stay, and sepsis were significantly increased in patients in the AA group ( p value < 0.05). There was no difference in the overall mortality between the two groups. AAs were associated with worse hospital course and complications but no difference in overall mortality rate. Age and cardiovascular disease seem to be predisposing risk factors for this. Further studies are needed to investigate potential strategies to prevent AAs development in this population.

Evidence of Advanced Pulmonary Vascular Remodeling in Obstructive Hypertrophic Cardiomyopathy With Pulmonary Hypertension.

BACKGROUND: Elevated mean pulmonary artery pressure (mPAP) is common in patients with hypertrophic cardiomyopathy (HCM) and heart failure symptoms. However, dynamic left ventricular (LV) outflow tract obstruction may confound interpretation of pulmonary hypertension (PH) pathophysiologic features in HCM when relying on resting invasive hemodynamic data alone. RESEARCH QUESTION: Do structural changes to the lung vasculature clarify PH pathophysiologic features in patients with HCM with progressive heart failure? STUDY DESIGN AND METHODS: Clinical data and ultrarare lung autopsy specimens were acquired retrospectively from the National Institutes of Health (1975-1992). Patients were included based on the availability of lung tissue and recorded mPAP. Discarded tissue from rejected lung donors served as control specimens. Histomorphology was performed on pulmonary arterioles and veins. Comparisons were calculated using the Student t test and Mann-Whitney U test; Pearson correlation was used to assess association between morphometric measurements and HCM cardiac and hemodynamic measurements. RESULTS: The HCM cohort (n = 7; mean ± SD age, 43 ± 18 years; 71% men) showed maximum mean ± SD LV wall thickness of 25 ± 2.8 mm, mean ± SD outflow tract gradient of 90 ± 30 mm Hg, median mPAP of 25 mm Hg (interquartile range [IQR], 6 mm Hg), median pulmonary artery wedge pressure (PAWP) of 16 mm Hg (IQR, 4 mm Hg), and median pulmonary vascular resistance of 1.8 Wood units (WU; IQR, 2.4 WU). Compared with control samples (n = 5), patients with HCM showed greater indexed pulmonary arterial hypertrophy (20.7 ± 7.2% vs 49.7 ± 12%; P < .001) and arterial wall fibrosis (11.5 ± 3.4 mm vs 21.0 ± 4.7 mm; P < .0001), which correlated with mPAP (r = 0.84; P = .018), PAWP (r = 0.74; P = .05), and LV outflow tract gradient (r = 0.78; P = .035). Compared with control samples, pulmonary vein thickness was increased by 2.9-fold (P = .008) in the HCM group, which correlated with mPAP (r = 0.81; P = .03) and LV outflow tract gradient (r = 0.83; P = .02). INTERPRETATION: To the best of our knowledge, these data demonstrate for the first time that in patients with obstructive HCM, heart failure is associated with pathogenic pulmonary vascular remodeling even when mPAP is elevated only mildly. These observations clarify PH pathophysiologic features in HCM, with future implications for clinical strategies that mitigate outflow tract obstruction.

Type I interferon-dependent IFIT3 signaling is critical for viral clearance in airway neutrophils.

Neutrophilic inflammation characterizes several respiratory viral infections including COVID-19-related ARDS, although its contribution to disease pathogenesis remains poorly understood. Here, we identified two neutrophil subpopulations (A1 and A2) in the airway compartment of 52 severe COVID-19 subjects, where loss of the A2 subset correlated with increased viral burden and reduced 30-days survival. A2 neutrophils showcased a discrete antiviral response with an increased interferon signature. Blockade of type I interferon attenuated viral clearance in A2 neutrophils and downregulated IFIT3 and key catabolic genes, demonstrating direct antiviral neutrophil function. Knockdown of IFIT3 in A2 neutrophils led to loss of IRF3 phosphorylation with consequent reduced viral catabolism, providing the first discrete mechanism of type I interferon signaling in neutrophils. The identification of this novel neutrophil phenotype and its association with severe COVID-19 outcomes emphasizes its likely importance in other respiratory viral infections and potential for new therapeutic approaches in viral illness.

IL-33 mediates Pseudomonas induced airway fibrogenesis and is associated with CLAD.

BACKGROUND: Long term outcomes of lung transplantation are impacted by the occurrence of chronic lung allograft dysfunction (CLAD). Recent evidence suggests a role for the lung microbiome in the occurrence of CLAD, but the exact mechanisms are not well defined. We hypothesize that the lung microbiome inhibits epithelial autophagic clearance of pro-fibrotic proteins in an IL-33 dependent manner, thereby augmenting fibrogenesis and risk for CLAD. METHODS: Autopsy derived CLAD and non-CLAD lungs were collected. IL-33, P62 and LC3 immunofluorescence was performed and assessed using confocal microscopy. Pseudomonas aeruginosa (PsA), Streptococcus Pneumoniae (SP), Prevotella Melaninogenica (PM), recombinant IL-33 or PsA-lipopolysaccharide was co-cultured with primary human bronchial epithelial cells (PBEC) and lung fibroblasts in the presence or absence of IL-33 blockade. Western blot analysis and quantitative reverse transcription (qRT) PCR was performed to evaluate IL-33 expression, autophagy, cytokines and fibroblast differentiation markers. These experiments were repeated after siRNA silencing and upregulation (plasmid vector) of Beclin-1. RESULTS: Human CLAD lungs demonstrated markedly increased expression of IL-33 and reduced basal autophagy compared to non-CLAD lungs. Exposure of co-cultured PBECs to PsA, SP induced IL-33, and inhibited PBEC autophagy, while PM elicited no significant response. Further, PsA exposure increased myofibroblast differentiation and collagen formation. IL-33 blockade in these co-cultures recovered Beclin-1, cellular autophagy and attenuated myofibroblast activation in a Beclin-1 dependent manner. CONCLUSION: CLAD is associated with increased airway IL-33 expression and reduced basal autophagy. PsA induces a fibrogenic response by inhibiting airway epithelial autophagy in an IL-33 dependent manner.

Interferon-dependent signaling is critical for viral clearance in airway neutrophils.

Neutrophilic inflammation characterizes several respiratory viral infections, including COVID-19-related acute respiratory distress syndrome, although its contribution to disease pathogenesis remains poorly understood. Blood and airway immune cells from 52 patients with severe COVID-19 were phenotyped by flow cytometry. Samples and clinical data were collected at 2 separate time points to assess changes during ICU stay. Blockade of type I interferon and interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) signaling was performed in vitro to determine their contribution to viral clearance in A2 neutrophils. We identified 2 neutrophil subpopulations (A1 and A2) in the airway compartment, where loss of the A2 subset correlated with increased viral burden and reduced 30-day survival. A2 neutrophils exhibited a discrete antiviral response with an increased interferon signature. Blockade of type I interferon attenuated viral clearance in A2 neutrophils and downregulated IFIT3 and key catabolic genes, demonstrating direct antiviral neutrophil function. Knockdown of IFIT3 in A2 neutrophils led to loss of IRF3 phosphorylation, with consequent reduced viral catabolism, providing the first discrete mechanism to our knowledge of type I interferon signaling in neutrophils. The identification of this neutrophil phenotype and its association with severe COVID-19 outcomes emphasizes its likely importance in other respiratory viral infections and potential for new therapeutic approaches in viral illness.

Primary Graft Dysfunction: The Role of Aging in Lung Ischemia-Reperfusion Injury.

Transplant centers around the world have been using extended criteria donors to remedy the ongoing demand for lung transplantation. With a rapidly aging population, older donors are increasingly considered. Donor age, at the same time has been linked to higher rates of lung ischemia reperfusion injury (IRI). This process of acute, sterile inflammation occurring upon reperfusion is a key driver of primary graft dysfunction (PGD) leading to inferior short- and long-term survival. Understanding and improving the condition of older lungs is thus critical to optimize outcomes. Notably, ex vivo lung perfusion (EVLP) seems to have the potential of reconditioning ischemic lungs through ex-vivo perfusing and ventilation. Here, we aim to delineate mechanisms driving lung IRI and review both experimental and clinical data on the effects of aging in augmenting the consequences of IRI and PGD in lung transplantation.

Aberrant Expression of ACO1 in Vasculatures Parallels Progression of Idiopathic Pulmonary Fibrosis.

Rationale: Idiopathic pulmonary fibrosis (IPF) is characterized by mitochondrial dysfunction. However, details about the non-mitochondrial enzymes that sustain the proliferative nature of IPF are unclear. Aconitases are a family of enzymes that sustain metabolism inside and outside mitochondria. It is hypothesized that aconitase 1 (ACO1) plays an important role in the pathogenesis of IPF given that ACO1 represents an important metabolic hub in the cytoplasm. Objectives: To determine if ACO1 expression in IPF lungs shows specific patterns that may be important in the pathogenesis of IPF. To determine the similarities and differences in ACO1 expression in IPF, bleomycin-treated, and aging lungs. Methods: ACO1 expression in IPF lungs were characterized and compared to non-IPF controls by western blotting, immunostaining, and enzymatic activity assay. ACO1-expressing cell types were identified by multicolor immunostaining. Using similar methods, the expression profiles of ACO1 in IPF lungs versus bleomycin-treated and aged mice were investigated. Measurements and main results: Lower lobes of IPF lungs, unlike non-IPF controls, exhibit significantly high levels of ACO1. Most of the signals colocalize with von Willebrand factor (vWF), a lineage marker for vascular endothelial cells. Bleomycin-treated lungs also show high ACO1 expressions. However, most of the signals colocalize with E-cadherin and/or prosurfactant protein C, representative epithelial cell markers, in remodeled areas. Conclusions: A characteristic ACO1 expression profile observed in IPF vasculatures may be a promising diagnostic target. It also may give clues as to how de novo angiogenesis contributes to the irreversible nature of IPF.

Single-cell transcriptome mapping identifies a local, innate B cell population driving chronic rejection after lung transplantation.

Bronchiolitis obliterans syndrome (BOS) is the main reason for poor outcomes after lung transplantation (LTx). We and others have recently identified B cells as major contributors to BOS after LTx. The extent of B cell heterogeneity and the relative contributions of B cell subpopulations to BOS, however, remain unclear. Here, we provide a comprehensive analysis of cell population changes and their gene expression patterns during chronic rejection after orthotopic LTx in mice. Of 11 major cell types, Mzb1-expressing plasma cells (PCs) were the most prominently increased population in BOS lungs. These findings were validated in 2 different cohorts of human BOS after LTx. A Bhlhe41, Cxcr3, and Itgb1 triple-positive B cell subset, also expressing classical markers of the innate-like B-1 B cell population, served as the progenitor pool for Mzb1+ PCs. This subset accounted for the increase in IgG2c production within BOS lung grafts. A genetic lack of Igs decreased BOS severity after LTx. In summary, we provide a detailed analysis of cell population changes during BOS. IgG+ PCs and their progenitors - an innate B cell subpopulation - are the major source of local Ab production and a significant contributor to BOS after LTx.

Pulmonary and Critical Care Considerations for e-Cigarette, or Vaping, Product Use-Associated Lung Injury.

BACKGROUND: In 2019, the United States experienced a nationwide outbreak of e-cigarette, or vaping, product use-associated lung injury (EVALI). More than one-half of these patients required admission to an ICU. RESEARCH QUESTION: What are the recent literature and expert opinions which inform the diagnosis and management of patients with critical illness with EVALI? STUDY DESIGN AND METHODS: To synthesize information critical to pulmonary/critical care specialists in the care of patients with EVALI, this study examined data available from patients hospitalized with EVALI between August 2019 and January 2020; reviewed the clinical course and critical care experience with those patients admitted to the ICU; and compiled opinion of national experts. RESULTS: Of the 2,708 patients with confirmed or probable EVALI requiring hospitalization as of January 21, 2020, a total of 1,604 (59.2%) had data available on ICU admission; of these, 705 (44.0%) were admitted to the ICU and are included in this analysis. The majority of ICU patients required respiratory support (88.5%) and in severe cases required intubation (36.1%) or extracorporeal membrane oxygenation (6.7%). The majority (93.0%) of these ICU patients survived to discharge. Review of the clinical course and expert opinion provided insight into: imaging; considerations for bronchoscopy; medical treatment, including use of empiric antibiotics, antiviral agents, and corticosteroids; respiratory support, including considerations for intubation, positioning maneuvers, and extracorporeal membrane oxygenation; and patient outcomes. INTERPRETATION: Review of the clinical course of patients with EVALI requiring ICU admission and compilation of expert opinion provided critical insight into pulmonary/critical care-specific considerations for this patient population. Because a large proportion of patients hospitalized with EVALI required ICU admission, it is important to remain prepared to care for patients with EVALI.

Spatial mapping of SARS-CoV-2 and H1N1 lung injury identifies differential transcriptional signatures.

Severe SARS-CoV-2 infection often leads to the development of acute respiratory distress syndrome (ARDS), with profound pulmonary patho-histological changes post-mortem. It is not clear whether ARDS from SARS-CoV-2 is similar to that observed in influenza H1N1, another common viral cause of lung injury. Here, we analyze specific ARDS regions of interest utilizing a spatial transcriptomic platform on autopsy-derived lung tissue from patients with SARS-CoV-2 (n = 3), H1N1 (n = 3), and a dual infected individual (n = 1). Enhanced gene signatures in alveolar epithelium, vascular tissue, and lung macrophages identify not only increased regional coagulopathy but also increased extracellular remodeling, alternative macrophage activation, and squamous metaplasia of type II pneumocytes in SARS-CoV-2. Both the H1N1 and dual-infected transcriptome demonstrated an enhanced antiviral response compared to SARS-CoV-2. Our results uncover regional transcriptional changes related to tissue damage/remodeling, altered cellular phenotype, and vascular injury active in SARS-CoV-2 and present therapeutic targets for COVID-19-related ARDS.

Recipient Age Impacts Long-Term Survival in Adult Subjects with Cystic Fibrosis after Lung Transplantation.

Rationale: Lung transplant is an effective treatment option providing survival benefit in patients with cystic fibrosis (CF). Several studies have suggested survival benefit in adults compared with pediatric patients with CF undergoing lung transplant. However, it remains unclear whether this age-related disparity persists in adult subjects with CF.Objectives: We investigated the impact of age at transplant on post-transplant outcomes in adult patients with CF.Methods: The United Network of Organ Sharing Registry was queried for all adult patients with CF who underwent lung transplantation between 1992 and 2016. Pertinent baseline characteristics, demographics, clinical parameters, and outcomes were recorded. The patients were divided into two groups based on age at transplant (18-29 yr old and 30 yr or older). The primary endpoint was survival time. Assessment of post-transplant survival was performed using Kaplan-Meier tests and log-rank tests with multivariable Cox proportional hazards analysis to adjust for confounding variables.Results: A total of 3,881 patients with CF underwent lung transplantation between 1992 and 2016; mean age was 31.0 (± 9.3) years. The 18-29-year-old at transplant cohort consisted of 2,002 subjects and the 30 years or older cohort had 1,879 subjects. Survival analysis demonstrated significantly higher survival in subjects in the 30 years or older cohort (9.47 yr; 95% confidence interval [CI], 8.7-10.2) compared with the 18-29-year-old cohort (5.21 yr; 95% CI, 4.6-5.8). After adjusting for confounders, survival remained higher in recipients aged 30 years or older (hazard ratio, 0.44; 95% CI, 0.2-0.9). Mortality due to allograft failure was significantly lower in patients with CF aged 30 years or older (28% vs. 36.5%; odds ratio [OR], 0.7; 95% CI, 0.6-0.8), whereas the incidence of malignancy was higher in the 30 years or older cohort (8% vs. 2.9%; OR, 3.0; 95% CI, 1.9-4.6).Conclusions: Age at transplant influences lung transplant outcomes in recipients with CF. Subjects with CF aged 30 years or older at transplant have superior survival compared with adult subjects with CF transplanted between the ages 18 and 29 years.

N-myc-interactor mediates microbiome induced epithelial to mesenchymal transition and is associated with chronic lung allograft dysfunction.

BACKGROUND: Recent evidence suggests a role for lung microbiome in occurrence of chronic lung allograft dysfunction (CLAD). However, the mechanisms linking the microbiome to CLAD are poorly delineated. We investigated a possible mechanism involved in microbial modulation of mucosal response leading to CLAD with the hypothesis that a Proteobacteria dominant lung microbiome would inhibit N-myc-interactor (NMI) expression and induce epithelial to mesenchymal transition (EMT). METHODS: Explant CLAD, non-CLAD, and healthy nontransplant lung tissue were collected, as well as bronchoalveolar lavage from 14 CLAD and matched non-CLAD subjects, which were followed by 16S rRNA amplicon sequencing and quantitative polymerase chain reaction (PCR) analysis. Pseudomonas aeruginosa (PsA) or PsA-lipopolysaccharide was cocultured with primary human bronchial epithelial cells (PBEC). Western blot analysis and quantitative reverse transcription (qRT) PCR was performed to evaluate NMI expression and EMT in explants and in PsA-exposed PBECs. These experiments were repeated after siRNA silencing and upregulation (plasmid vector) of EMT regulator NMI. RESULTS: 16S rRNA amplicon analyses revealed that CLAD patients have a higher abundance of phyla Proteobacteria and reduced abundance of the phyla Bacteroidetes. At the genera level, CLAD subjects had an increased abundance of genera Pseudomonas and reduced Prevotella. Human CLAD airway cells showed a downregulation of the N-myc-interactor gene and presence of EMT. Furthermore, exposure of human primary bronchial epithelial cells to PsA resulted in downregulation of NMI and induction of an EMT phenotype while NMI upregulation resulted in attenuation of this PsA-induced EMT response. CONCLUSIONS: CLAD is associated with increased bacterial biomass and a Proteobacteria enriched airway microbiome and EMT. Proteobacteria such as PsA induces EMT in human bronchial epithelial cells via NMI, demonstrating a newly uncovered mechanism by which the microbiome induces cellular metaplasia.

Prolyl endopeptidase contributes to early neutrophilic inflammation in acute myocardial transplant rejection.

Altered inflammation and tissue remodeling are cardinal features of cardiovascular disease and cardiac transplant rejection. Neutrophils have increasingly been understood to play a critical role in acute rejection and early allograft failure; however, discrete mechanisms that drive this damage remain poorly understood. Herein, we demonstrate that early acute cardiac rejection increases allograft prolyl endopeptidase (PE) in association with de novo production of the neutrophil proinflammatory matrikine proline-glycine-proline (PGP). In a heterotopic murine heart transplant model, PGP production and PE activity were associated with early neutrophil allograft invasion and allograft failure. Pharmacologic inhibition of PE with Z-Pro-prolinal reduced PGP, attenuated early neutrophil graft invasion, and reduced proinflammatory cytokine expression. Importantly, these changes helped preserve allograft rejection-free survival and function. Notably, within 2 independent patient cohorts, both PGP and PE activity were increased among patients with biopsy-proven rejection. The observed induction of PE and matrikine generation provide a link between neutrophilic inflammation and cardiovascular injury, represent a potential target to reduce allogenic immune responses, and uncover a mechanism of cardiovascular disease that has been previously unrecognized to our knowledge.