Endothelial-to-Mesenchymal Transition in Human and Murine Models of Congenital Diaphragmatic Hernia

<b><i>Introduction:</i></b> Congenital diaphragmatic hernia (CDH) is a complex congenital disorder, characterized by pulmonary hypertension (PH) and hypoplasia. PH secondary to CDH (CDH-PH) features devastating morbidity and mortality (25–30%) among neonates. An unmet need is determining mechanisms triggering CDH-PH to save infants. Prior data suggest abnormal remodeling of the pulmonary vascular extracellular matrix (ECM), presumed to be driven by endothelial-to-mesenchymal transition (EndoMT), hinders postnatal vasodilation and limits anti-PH therapy in CDH. There are limited data on the role of EndoMT in CDH-PH. <b><i>Methods:</i></b> The purpose of the study was to investigate how EndoMT contributes to CDH-PH by identifying cells undergoing EndoMT noted by alpha smooth muscle actin (α-SMA) expression in human umbilical vein endothelial cells (HUVECs) and lung tissue obtained from murine pups using the nitrofen model. <i>N</i> = 8 CDH, <i>N</i> = 8 control HUVECs were stained for α-SMA and CD31 after being exposed for 24 h to TGFB, a known EndoMT promoter. <i>N</i> = 8 nitrofen, <i>N</i> = 8 control murine pup lungs were also stained for α-SMA and CD31. α-SMA and CD31 expression was quantified in HUVECs and murine tissue using Fiji imaging software and normalized to the total number of cells per slide noted by DAPI staining. <b><i>Results:</i></b> CDH HUVECs demonstrated a 1.1-fold increase in α-SMA expression (<i>p</i> = 0.02). The murine model did not show statistical significance between nitrofen and control pup lungs; however, there was a 0.4-fold increase in α-SMA expression with a 0.8-fold decrease in CD31 expression in the nitrofen pup lungs when compared to controls. <b><i>Conclusion:</i></b> These results suggest that EndoMT could potentially play a role in the ECM remodeling seen in CDH-PH.