Abstract
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 β2 (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.
Abstract
Components of the renin–angiotensin system are increased in chronic lung allograft dysfunction (CLAD) fibrosis. Angiotensin II-regulated proteins in bronchoalveolar lavage identify concurrent and predict future CLAD in lung transplant recipients. https://bit.ly/3eRYez7
Footnotes
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Author contributions: G. Berra: clinical screening, cohort design, sample collection and organisation, immunostaining, qPCR, Western blotting, data analysis, writing of manuscript. S. Farkona: experimental design, mass spectrometry, Western blotting, data analysis, writing of manuscript. Z. Mohammed-Ali: mass spectrometry. M. Kotlyar: bioinformatic analyses, machine learning algorithms, help with manuscript writing. L. Levy: cohort design, collection and handling of samples, help with manuscript writing. S. Clotet-Freixas: Western blotting. P. Ly: immunostaining. B. Renaud-Picard: clinical screening, collection of samples, histological grading. G. Zehong: help with immunostaining protocols. T. Daigneault: collection of samples. A. Duong: collection and organisation of samples. I. Batruch: technical assistance with mass spectrometry. I. Jurisica: protein interactome generation, bioinformatic analyses supervision. A. Konvalinka and T. Martinu: conception of the research question, cohort design, experimental design, supervision of experiments, data analysis, manuscript writing and editing.
Conflict of interest: G. Berra has nothing to disclose.
Conflict of interest: S. Farkona has nothing to disclose.
Conflict of interest: Z. Mohammed-Ali has nothing to disclose.
Conflict of interest: M. Kotlyar has nothing to disclose.
Conflict of interest: L. Levy has nothing to disclose.
Conflict of interest: S. Clotet-Freixas has nothing to disclose.
Conflict of interest: P. Ly has nothing to disclose.
Conflict of interest: B. Renaud-Picard has nothing to disclose.
Conflict of interest: G. Zehong has nothing to disclose.
Conflict of interest: T. Daigneault has nothing to disclose.
Conflict of interest: A. Duong has nothing to disclose.
Conflict of interest: I. Batruch has nothing to disclose.
Conflict of interest: I. Jurisica reports grants and nonfinancial support (in-kind contribution to grants) from IBM, personal fees for lectures and other (reimbursement of expenses) from Novartis and Canadian Rheumatology Association, outside the submitted work.
Conflict of interest: A. Konvalinka has nothing to disclose.
Conflict of interest: T. Martinu reports grants from Canadian Donation and Transplantation Research Program and Di Pochi funds, during the conduct of the study; grants from Sanofi, nonfinancial support from APCBio, outside the submitted work.
Support statement: The authors disclose receipt of the following financial support for this research: Canadian National Transplant Research Program, Kidney Foundation of Canada Predictive Biomarker Grant, Canadian Institute for Health Research (CIHR), Kidney Research Scientist Core Education, National Training (KRESCENT) program, Di Pochi funds, Toronto General and Western Hospital Foundation, University Health Network Multi Organ Transplant Program, and Les Hôpitaux Universitaires de Genève. Computational analyses were supported in part by grants from the Ontario Research Fund (34876) and Canada Foundation for Innovation (29272, 225404 and 33536). Funding information for this article has been deposited with the Crossref Funder Registry.
- Received July 30, 2020.
- Accepted March 6, 2021.
- Copyright ©The authors 2021. For reproduction rights and permissions contact permissions{at}ersnet.org