A multidimensional classifier to support lung transplant referral in patients with pulmonary fibrosis.

BACKGROUND: Lung transplantation remains the sole curative option for patients with idiopathic pulmonary fibrosis (IPF), but donor organs remain scarce, and many eligible patients die before transplant. Tools to optimize the timing of transplant referrals are urgently needed. METHODS: Least absolute shrinkage and selection operator was applied to clinical and proteomic data generated as part of a prospective cohort study of interstitial lung disease (ILD) to derive clinical, proteomic, and multidimensional logit models of near-term death or lung transplant within 18 months of blood draw. Model-fitted values were dichotomized at the point of maximal sensitivity and specificity, and decision curve analysis was used to select the best-performing classifier. We then applied this classifier to independent IPF and non-IPF ILD cohorts to determine test performance characteristics. Cohorts were restricted to patients aged ≤72 years with body mass index 18 to 32 to increase the likelihood of transplant eligibility. RESULTS: IPF derivation, IPF validation, and non-IPF ILD validation cohorts consisted of 314, 105, and 295 patients, respectively. A multidimensional model comprising 2 clinical variables and 20 proteins outperformed stand-alone clinical and proteomic models. Following dichotomization, the multidimensional classifier predicted near-term outcome with 70% sensitivity and 92% specificity in the IPF validation cohort and 70% sensitivity and 80% specificity in the non-IPF ILD validation cohort. CONCLUSIONS: A multidimensional classifier of near-term outcomes accurately discriminated this end-point with good test performance across independent IPF and non-IPF ILD cohorts. These findings support refinement and prospective validation of this classifier in transplant-eligible individuals.

Genetic deficiency of the transcription factor NFAT1 confers protection against fibrogenic responses independent of immune influx.

Idiopathic pulmonary fibrosis (IPF) is marked by unremitting matrix deposition and architectural distortion. Multiple profibrotic pathways contribute to the persistent activation of mesenchymal cells (MCs) in fibrosis, highlighting the need to identify and target common signaling pathways. The transcription factor nuclear factor of activated T cells 1 (NFAT1) lies downstream of second messenger calcium signaling and has been recently shown to regulate key profibrotic mediator autotaxin (ATX) in lung MCs. Herein, we investigate the role of NFAT1 in regulating fibroproliferative responses during the development of lung fibrosis. Nfat1-/--deficient mice subjected to bleomycin injury demonstrated improved survival and protection from lung fibrosis and collagen deposition as compared with bleomycin-injured wild-type (WT) mice. Chimera mice, generated by reconstituting bone marrow cells from WT or Nfat1-/- mice into irradiated WT mice (WT→WT and Nfat1-/-→WT), demonstrated no difference in bleomycin-induced fibrosis, suggesting immune influx-independent fibroprotection in Nfat1-/- mice. Examination of lung tissue and flow sorted lineageneg/platelet-derived growth factor receptor alpha (PDGFRα)pos MCs demonstrated decreased MC numbers, proliferation [↓ cyclin D1 and 5-ethynyl-2'-deoxyuridine (EdU) incorporation], myofibroblast differentiation [↓ α-smooth muscle actin (α-SMA)], and survival (↓ Birc5) in Nfat1-/- mice. Nfat1 deficiency abrogated ATX expression in response to bleomycin in vivo and MCs derived from Nfat1-/- mice demonstrated decreased ATX expression and migration in vitro. Human IPF MCs demonstrated constitutive NFAT1 activation, and regulation of ATX in these cells by NFAT1 was confirmed using pharmacological and genetic inhibition. Our findings identify NFAT1 as a critical mediator of profibrotic processes, contributing to dysregulated lung remodeling and suggest its targeting in MCs as a potential therapeutic strategy in IPF.NEW & NOTEWORTHY Idiopathic pulmonary fibrosis (IPF) is a fatal disease with hallmarks of fibroblastic foci and exuberant matrix deposition, unknown etiology, and ineffective therapies. Several profibrotic/proinflammatory pathways are implicated in accelerating tissue remodeling toward a honeycombed end-stage disease. NFAT1 is a transcriptional factor activated in IPF tissues. Nfat1-deficient mice subjected to chronic injury are protected against fibrosis independent of immune influxes, with suppression of profibrotic mesenchymal phenotypes including proliferation, differentiation, resistance to apoptosis, and autotaxin-related migration.

A 30-Year-Old Immune Deficient Woman With Persistent Cough and Shortness of Breath.

CASE PRESENTATION: A 30-year-old woman was referred with increasing shortness of breath and cough in the setting of GATA2 deficiency. She initially presented 9 years previously with recurrent episodes of pneumonia and sinusitis. Genetic testing revealed a heterozygous GATA2 mutation (c.988C>T). She has since had multiple infections that have included necrotizing fasciitis of the right thumb, recurrent pilonidal infections (which required 23 procedures), esophageal candidiasis, and human papillomavirus-positive high-grade squamous intraepithelial lesion of the cervix. Serial bone marrow biopsy specimens showed persistent hypocellularity (20% to 60%) with intermittent erythroid atypia and variable detection of trisomy 8, which were concerning for evolving myelodysplastic syndrome. One year before the current admission, she was diagnosed with disseminated Mycobacterium avium complex and was treated with rifabutin, ethambutol, and azithromycin. She was taking voriconazole, acyclovir, and trimethoprim-sulfamethoxazole prophylaxis.

Lung microbiota predict chronic rejection in healthy lung transplant recipients: a prospective cohort study.

BACKGROUND: Alterations in the respiratory microbiome are common in chronic lung diseases, correlate with decreased lung function, and have been associated with disease progression. The clinical significance of changes in the respiratory microbiome after lung transplant, specifically those related to development of chronic lung allograft dysfunction (CLAD), are unknown. The aim of this study was to evaluate the effect of lung microbiome characteristics in healthy lung transplant recipients on subsequent CLAD-free survival. METHODS: We prospectively studied a cohort of lung transplant recipients at the University of Michigan (Ann Arbor, MI, USA). We analysed characteristics of the respiratory microbiome in acellular bronchoalveolar lavage fluid (BALF) collected from asymptomatic patients during per-protocol surveillance bronchoscopy 1 year after lung transplantation. For our primary endpoint, we evaluated a composite of development of CLAD or death at 500 days after the 1-year surveillance bronchoscopy. Our primary microbiome predictor variables were bacterial DNA burden (total 16S rRNA gene copies per mL of BALF, quantified via droplet digital PCR) and bacterial community composition (determined by bacterial 16S rRNA gene sequencing). Patients' lung function was followed serially at least every 3 months by spirometry, and CLAD was diagnosed according to International Society of Heart and Lung Transplant 2019 guidelines. FINDINGS: We analysed BALF from 134 patients, collected during 1-year post-transplant surveillance bronchoscopy between Oct 21, 2005, and Aug 25, 2017. Within 500 days of follow-up from the time of BALF sampling, 24 (18%) patients developed CLAD, five (4%) died before confirmed development of CLAD, and 105 (78%) patients remained CLAD-free with complete follow-up. Lung bacterial burden was predictive of CLAD development or death within 500 days of the surveillance bronchoscopy, after controlling for demographic and clinical factors, including immunosuppression and bacterial culture results, in a multivariable survival model. This relationship was evident when burden was analysed as a continuous variable (per log10 increase in burden, HR 2·49 [95% CI 1·38-4·48], p=0·0024) or by tertiles (middle vs lowest bacterial burden tertile, HR 4·94 [1·25-19·42], p=0·022; and highest vs lowest, HR 10·56 [2·53-44·08], p=0·0012). In patients who developed CLAD or died, composition of the lung bacterial community significantly differed to that in patients who survived and remained CLAD-free (on permutational multivariate analysis of variance, p=0·047 at the taxonomic level of family), although differences in community composition were associated with bacterial burden. No individual bacterial taxa were definitively associated with CLAD development or death. INTERPRETATION: Among asymptomatic lung transplant recipients at 1-year post-transplant, increased lung bacterial burden is predictive of chronic rejection and death. The lung microbiome represents an understudied and potentially modifiable risk factor for lung allograft dysfunction. FUNDING: US National Institutes of Health, Cystic Fibrosis Foundation, Brian and Mary Campbell and Elizabeth Campbell Carr research gift fund.

Interleukin 6 trans-signaling is a critical driver of lung allograft fibrosis.

Histopathologic examination of lungs afflicted by chronic lung allograft dysfunction (CLAD) consistently shows both mononuclear cell (MNC) inflammation and mesenchymal cell (MC) fibroproliferation. We hypothesize that interleukin 6 (IL-6) trans-signaling may be a critical mediator of MNC-MC crosstalk and necessary for the pathogenesis of CLAD. Bronchoalveolar lavage (BAL) fluid obtained after the diagnosis of CLAD has approximately twofold higher IL-6 and soluble IL-6 receptor (sIL-6R) levels compared to matched pre-CLAD samples. Human BAL-derived MCs do not respond to treatment with IL-6 alone but have rapid and prolonged JAK2-mediated STAT3 Tyr705 phosphorylation when exposed to the combination of IL-6 and sIL-6R. STAT3 phosphorylation within MCs upregulates numerous genes causing increased invasion and fibrotic differentiation. MNC, a key source of both IL-6 and sIL-6R, produce minimal amounts of these proteins at baseline but significantly upregulate production when cocultured with MCs. Finally, the use of an IL-6 deficient recipient in a murine orthotopic transplant model of CLAD reduces allograft fibrosis by over 50%. Taken together these results support a mechanism where infiltrating MNCs are stimulated by resident MCs to release large quantities of IL-6 and sIL-6R which then feedback onto the MCs to increase invasion and fibrotic differentiation.

Fibroproliferation in chronic lung allograft dysfunction: Association of mesenchymal cells in bronchoalveolar lavage with phenotypes and survival.

BACKGROUND: Chronic lung allograft dysfunction (CLAD), the primary cause of poor outcome after lung transplantation, arises from fibrotic remodeling of the allograft and presents as diverse clinical phenotypes with variable courses. Here, we investigate whether bronchoalveolar lavage (BAL) mesenchymal cell activity at CLAD onset can inform regarding disease phenotype, progression, and survival. METHODS: Mesenchymal cell colony-forming units (CFUs) were measured in BAL obtained at CLAD onset (n = 77) and CLAD-free time post-transplant matched controls (n = 77). CFU counts were compared using Wilcoxon's rank-sum test. Cox proportional hazards and restricted means models were utilized to investigate post-CLAD survival. RESULTS: Higher mesenchymal CFU counts were noted in BAL at the time of CLAD onset than in CLAD-free controls. Patients with restrictive allograft syndrome had higher BAL mesenchymal CFU count at CLAD onset than patients with bronchiolitis obliterans syndrome (p = 0.011). Patients with high mesenchymal CFU counts (≥10) at CLAD onset had worse outcomes than those with low (<10) CFU counts, with shorter average survival (2.64 years vs 4.25 years; p = 0.027) and shorter progression-free survival, defined as time to developing either CLAD Stage 3 or death (0.97 years vs 2.70 years; p < 0.001). High CFU count remained predictive of decreased overall survival and progression-free survival after accounting for the CLAD phenotype and other clinical factors in multivariable analysis. CONCLUSIONS: Fulminant fibroproliferation with higher mesenchymal CFU counts in BAL is noted in restrictive allograft syndrome and is independently associated with poor survival after CLAD onset.