Phospholipid Profile of Amniotic Fluid in Ovine Model of Congenital Diaphragmatic Hernia (CDH): The Effect of Fetal Tracheal Occlusion Maria Chiara Mimmi,* ,† Maurizio Ballico, † Francesco Amoroso, † Valeria Calcaterra, ‡ Mario Marotta, §,∥ Jose Luis Peiro, §,∥ and Gloria Pelizzo ⊥ † Department of Medical and Biological Sciences, University of Udine, Piazzale M. Kolbe 4, 33100 Udine, Italy ‡ Department of the Mother and Child Health, Pediatric Unit, IRCCS Policlinico San Matteo Foundation Pavia and Department of Internal Medicine, University of Pavia, Viale Camillo Golgi 19, 27100 Pavia, Italy § Pediatric Surgery, Orthopaedics and Bioengineering Laboratory, VHIR Research Institute, Vall d’Hebron University Hospital, Passeig de la Vall d'Hebron, 119, 08035 Barcelona, Spain ∥ Center for Fetal, Cellular and Molecular Therapy, Cincinnati Children’s Hospital Medical Center (CCHMC), 3333 Burnet Avenue, Cincinnati, Ohio 45229, United States ⊥ Department of the Mother and Child Health, Pediatric Surgery Unit, IRCCS Policlinico San Matteo Foundation Pavia and University of Pavia, Viale Camillo Golgi 19, 27100 Pavia, Italy * S Supporting Information ABSTRACT: Fetal endoscopic tracheal occlusion has been proposed as a prenatal intervention to ameliorate congenital diaphragmatic hernia (CDH) prognosis. Tracheal occlusion (TO) prevents pulmonary fluid egress, leading to tissue expansion, reversal of lung hypoplasia, and potential maturation. Fetal lung maturity strongly correlates with amniotic fluid (AF) phospholipidic composition. In this preliminary study, we characterized the AF phospholipidic profile in CDH-induced, TO-treated, and healthy fetal lambs to define the prenatal treatment benefits of TO on lung maturity. CDH induction was performed at 70 days of gestation, TO was carried out at 102 days of gestation, and caesarean section was carried out at 136 days of gestation. AF samples, taken at 102−136 days of gestation, were evaluated using mass spectrometry. The analysis focused on phosphatidylcholines (PCs) and sphingomyelins (SMs). The most abundant phosphatidylcholine species retrieved in healthy AF was POPC [PC(18:1/16:0)], while the level of DPPC [PC(16:0/16:0)] was extremely low at both gestational ages. CDH induction caused a decrease in POPC and many other PCs. A substantial return of some PCs, in particular POPC, PC(34:2) and PC(18:0/16:0), to a more physiological level was prompted by TO. SMs were unaltered. The AF phospholipidic profile could provide prenatal prognostic markers of CDH and possible indices of lung maturation after fetal treatment. KEYWORDS: lipidomics, lung surfactant, phospholipids, phosphatidylcholine, LC−MS, fetal lung maturity, biomarkers, amniotic fluid, ovine models, congenital diaphragmatic hernia ■ INTRODUCTION CDH affects 1 in 2000 to 3000 newborns. It is considered a life- threatening malformation because it causes lung hypoplasia and pulmonary hypertension. Despite recent progress in prenatal and neonatal care, the mortality rate remains high (67%). Morbidity in surving neonates is also observed in more than 50% of cases, including pulmonary abnormalities in as many as 50%, gastroesophageal reflux disease in 20−72%, failure to thrive in 20−30%, skeletal abnormalities in 16−48%, and neurological deficits in 10−30%. 1 Postnatal surgical repair is the treatment of choice (more than 95%) in patients with a favorable prognosis. 2 Prenatal therapy, including fetal endoscopic tracheal occlusion (FETO), has been introduced in severe CDH with a poor prognosis to improve the outcome. Fetal treatment promotes lung growth through increased intraluminal pres- sure. 3 Endotracheal fluid, a major determinant of fetal lung growth, 4 is normally exchanged with amniotic fluid through the physiological mechanism of swallowing and alveolar lavage. It is plausible that the accelerated lung maturation determined by tracheal occlusion is due to not only a biophysical effect but also the temporary trapping of endotracheal fluid in the lungs. The assessment of fetal lung maturity (FLM) is a key issue for the clinical care of preterm and lung disorder-affected infants, as well as for decisions regarding the timing of delivery Received: October 29, 2014 Article pubs.acs.org/jpr © XXXX American Chemical Society A DOI: 10.1021/pr501120x J. Proteome Res. XXXX, XXX, XXX−XXX