Pathophysiological studies in neonatal tracheal aspirate derived mesenchymal stromal cells and animal models to determine the role of Pbx1 and other candidate genes in congenital diaphragmatic hernia.
Final Report Abstract
Congenital diaphragmatic hernia (CDH) affects 1 in 2500 newborns. lt is characterized by underdeveloped lungs and a congenital defect in the diaphragm. Despite advances in intensive medical and surgical therapy, up to 30% of patients still succumb to lung malformations, and survivors often suffer from lifelong complications. Our limited understanding of the pathophysiology hinders the development of innovative therapeutic approaches. An established animal model, the Nitrofen rat model, is used for experimental investigation of CDH and has revealed changes in specific molecular markers in the hypoplastic CDH lungs. However, to date, these findings have only been partially applicable to the human situation. In this study, we cumulatively demonstrated that pro-inflammatory signaling pathways are particularly enriched in abnormal lung development in the animal model and in CDH in humans. Targeted blockade of these pathologically altered pathways led to the correction of impaired lung growth. To do this, we initially established a new method for isolating epithelial lung stem cells from routinely collected tracheal aspirates of newborns and investigated them in culture using functional and descriptive assays. In epithelial precursor cells from newborns with CDH, we identified transcriptional (bulk RNA sequencing) and epigenetic profiles {ATAC sequencing) indicating pro-inflammatory changes, including increased activity of the transcription factor NF-kB. Inhibition of NF-kB using glucocorticoids and specific NF-kB antagonists corrected the typical CDH phenotype in both human cell lines and the animal model. In a previous study using the established Nitrofen rat model for CDH, we identified a comparable pro-inflammatory program during later lung development in hypoplastic lungs through proteomic analyses (untargeted proteomics). The results from our in vitro studies in humans and in vivo studies in the animal model were validated through immunostaining in human fetal lung tissue from CDH patients and controls. In summary, we have shown in various translational models that a pro-inflammatory molecular program is active in hypoplastic CDH lungs, which could serve as a target for new therapeutic approaches to treat lung hypoplasia in CDH.
Publications
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Administration of amniotic fluid stem cell extracellular vesicles promotes development of fetal hypoplastic lungs by immunomodulating lung macrophages. Cold Spring Harbor Laboratory.
Antounians, Lina; Figueira, Rebeca Lopes; Kukreja, Bharti; Zani-Ruttenstock, Elke; Khalaj, Kasra; Montalva, Louise; Doktor, Fabian; Obed, Mikal; Blundell, Matisse; Wu, Taiyi; Chan, Cadia; Wagner, Richard; Lacher, Martin; Wilson, Michael D.; Kalish, Brian T. & Zani, Augusto
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Primary culture of tracheal aspirate-derived human airway basal stem cells. STAR Protocols, 3(2), 101390.
Amonkar, Gaurang M.; Wagner, Richard; Bankoti, Kamakshi; Shui, Jessica E.; Ai, Xingbin & Lerou, Paul H.
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Proteomic Profiling of Hypoplastic Lungs Suggests an Underlying Inflammatory Response in the Pathogenesis of Abnormal Lung Development in Congenital Diaphragmatic Hernia. Annals of Surgery, 278(2), e411-e421.
Wagner, Richard; Lieckfeldt, Paula; Piyadasa, Hadeesha; Markel, Moritz; Riedel, Jan; Stefanovici, Camelia; Peukert, Nicole; Patel, Daywin; Derraugh, Gabrielle; Min, Suyin A. Lum; Gosemann, Jan-Hendrik; Deprest, Jan; Pascoe, Christopher D.; Tse, Wai Hei; Lacher, Martin; Mookherjee, Neeloffer & Keijzer, Richard
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A Tracheal Aspirate-derived Airway Basal Cell Model Reveals a Proinflammatory Epithelial Defect in Congenital Diaphragmatic Hernia. American Journal of Respiratory and Critical Care Medicine, 207(9), 1214-1226.
Wagner, Richard; Amonkar, Gaurang M.; Wang, Wei; Shui, Jessica E.; Bankoti, Kamakshi; Tse, Wai Hei; High, Frances A.; Zalieckas, Jill M.; Buchmiller, Terry L.; Zani, Augusto; Keijzer, Richard; Donahoe, Patricia K.; Lerou, Paul H. & Ai, Xingbin
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Overactivated epithelial NF-κB disrupts lung development in human and nitrofen CDH. Cold Spring Harbor Laboratory.
Dylong, Florentine; Riedel, Jan; Amonkar, Gaurang M.; Peukert, Nicole; Lieckfeldt, Paula; Sturm, Katinka; Höxter, Benedikt; Tse, Wai Hei; Miyake, Yuichiro; Mayer, Steffi; Keijzer, Richard; Lacher, Martin; Ai, Xingbin; Gosemann, Jan-Hendrik & Wagner, Richard
