Abstract 18338: TBX4 Loss-of-Function Within the Embryonic Lung Mesenchyme Disrupts Postnatal Lung Development and Increases Susceptibility to Pulmonary Hypertension

Abstract

Introduction: The T-box 4 (TBX4) transcription factor is recognized as a risk gene in pulmonary arterial hypertension (PAH). TBX4 is expressed within the lung mesenchyme in the developing lung and is required for lung branching morphogenesis. Genetic studies report that PAH cases with TBX4 mutations display an earlier onset of the disease. In addition, studies have reported carriers of TBX4 mutations with diffused alveolar development and pulmonary hypertension (PH). Currently, there is a lack of evidence on the effect of TBX4 loss-of-function (LoF) on later stages of lung development and predisposition to lung disease. Hypothesis: Lung mesenchyme Tbx4 deletion within the developing lung disrupts essential gene regulatory networks leading to abnormal postnatal lung development and susceptibility to pulmonary hypertension. Methods: We have generated Tbx4 conditional knockout ( Tbx4-CKO ) mice in which Cre recombinase removes the exon 5 of Tbx4 within the embryonic lung mesenchyme, creating a null allele. To examine alveolarization, we harvested postnatal day 14 (P14) and 36 (P36) lungs and calculated mean linear intercept (MLI). RNA-seq was performed at both stages to identify disrupted gene networks in Tbx4-CKO lungs. RVSP was measured by right heart catheterization in six-month-old mice to evaluate for PH. Results: Tbx4-CKO lungs show increasing alveolar simplification at P14 and P36, as confirmed by increases in MLI of 24.8% and 34.4%, respectively. Tbx4-CKO mice show a statistically significant increase of 10.4% in RVSP, and preliminary analysis of vessel wall thickness suggests mild vascular remodeling in their lungs. RNA-seq analysis shows enrichment of pathways (canonical WNT, VEGF, and BMP signaling) and genes ( Lgr5 , Tnc , Wnt3a , Areg , Gdf2 , and Bmper ) relevant to lung alveologenesis, angiogenesis, and PH. Conclusions: TBX4 LoF affects lung alveolar development, similar to observations made in human cases with TBX4 mutations. The clinical presentation of such cases is broad, but our mice show features of bronchopulmonary dysplasia, including alveolar simplification and PH. Further analysis of our RNA-seq data and planned single-cell analyses should provide insight into the role of TBX4 in alveolarization and PH risk.