Impaired lung function secondary to fibrotic scarring and obliteration of small airways, termed bronchiolitis obliterans syndrome (BOS), is the major cause of chronic graft failure and mortality following lung transplantation. Understanding mechanistic pathways involved in lung allograft fibrogenesis is the key to novel therapeutic approaches in this arena. Studies of mesenchymal cells (MCs) from human lung allografts have led us to target a novel signaling pathway of MC mobilization and activation. An increase in number of donor-derived MCs is seen in the bronchoalveolar lavage (BAL) of patients with BOS. MCs from BOS lungs also demonstrated a stable activated phenotype marked by an increased collagen and ?-catenin protein expression as well as an increased autotaxin (ATX) secretory activity. ATX is a secreted lysophopholipase D which generates lysophosphatidic acid (LPA), a bioactive lipid with a role in tissue fibrosis, from membrane lipids. Our published work has revealed a unique signaling cascade where through LPA1 ligation and subsequent phosphokinase-C mediated glycogen synthase kinase-3? activation, LPA leads to ?-catenin stabilization and transcriptional activation. New preliminary data demonstrates that autocrine ATX secretion and LPA1 ligation contributes to stable ?-catenin and collagen upregulation noted in MCs isolated from human fibrotic lung grafts. Furthermore, LPA levels were noted to be higher in BAL samples from patients with BOS. This human data has led us to propose experiments investigating the in vivo role of ATX-LPA-?-catenin pathway in pathogenesis of BO and to investigate if LPA1 antagonism can be a potential therapeutic option in this disease. To achieve these aims we have established a whole lung mouse orthotopic lung transplant model where a moderate MHC mismatch leads to development of airway fibrosis by day 28.
In Aim 1, we propose to investigate the time course of ATX and LPA upregulation in the resident MCs and other cellular components of the allograft in vivo using this murine model. We will utilize mice with BAC florescent labeling of Foxf1, a transcription factor seen in embryonic lung mesenchyme and shown to be highly expressed in lung MCs, to specifically identify and study graft-resident MCs. These studies will also inform us of the role of graft-resident MCs in fibrogenesis.
Aim 2 will focus on understanding the role of ?-catenin in MC differentiation and dissecting the mechanisms by which it affects the cells fibrotic functions. The mechanistic pathways will be studied in human cells. However, we will utilize a reporter mouse (Axin2LacZ) to follow ?-catenin transcriptional activity during allograft fibrogenesis and test the in vivo rol of LPA1 ligation in MC ?-catenin activation.
In Aim 3 we will determine if LPA levels in BAL samples can predict subsequent development of BOS. Importantly, we will test if LPA1 antagonist can prevent onset or progression of allograft fibrosis in the murine orthotopic single lung transplant model. The proposed work will be the first investigation of ATX/LPA/LPA1/ ?-catenin signaling in BOS and will provide important rationale for future clinical trials targeting this pathway in lung transplantation.

Public Health Relevance

This study will investigate mechanism(s) of development of chronic graft failure in lung transplant recipients. Lung transplant has the lowest long term survival among all solid organ transplants and this is primarily secondary to development of airway fibrosis leading to graft failure. We propose a novel investigation of an important lipid pathway in the cells derived directly from the lung washings of transplant recipients. These studies will be combined with animal studies of these signaling molecules and their inhibition in a model of airway rejection. Our work is therefore expected to not only provide novel information regarding mechanism of fibrosis but also provide direct rationale for future human studies investigating drug targeting this pathway in lung transplant recipients.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Lung Injury, Repair, and Remodeling Study Section (LIRR)
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Eu, Jerry Pc
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University of Michigan Ann Arbor
Internal Medicine/Medicine
Schools of Medicine
Ann Arbor
United States
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Belloli, Elizabeth A; Lama, Vibha N (2016) Spirometry States the Obvious: Recognizing Bronchiolitis Obliterans Syndrome Early after Hematopoietic Cell Transplantation. Ann Am Thorac Soc 13:1883-1884
Walker, Natalie M; Belloli, Elizabeth A; Stuckey, Linda et al. (2016) Mechanistic Target of Rapamycin Complex 1 (mTORC1) and mTORC2 as Key Signaling Intermediates in Mesenchymal Cell Activation. J Biol Chem 291:6262-71
Mimura, Takeshi; Walker, Natalie; Aoki, Yoshiro et al. (2015) Local origin of mesenchymal cells in a murine orthotopic lung transplantation model of bronchiolitis obliterans. Am J Pathol 185:1564-74
Belloli, Elizabeth A; Wang, Xin; Murray, Susan et al. (2015) Longitudinal Forced Vital Capacity Monitoring as a Prognostic Adjunct after Lung Transplantation. Am J Respir Crit Care Med 192:209-18
Shah, R J; Diamond, J M; Cantu, E et al. (2015) Objective Estimates Improve Risk Stratification for Primary Graft Dysfunction after Lung Transplantation. Am J Transplant 15:2188-96
Dickson, Robert P; Erb-Downward, John R; Prescott, Hallie C et al. (2015) Intraalveolar Catecholamines and the Human Lung Microbiome. Am J Respir Crit Care Med 192:257-9