Analysis of Differentially Regulated Genes in Lung Tissue

Pulmonary arterial hypertension (PAH) is a progressive fatal disease that is characterized by pathological pulmonary artery remodeling, in which endothelial cell (EC) dysfunction is critically involved. We herein describe a previously unknown role of endothelial angiocrine in pulmonary hypertension. By searching for genes highly expressed in lung microvascular ECs, we identified inhibin--A (INHBA) as an angiocrine factor produced by pulmonary capillaries. We found that excess production of INHBA by ECs impairs the endothelial function in an autocrine manner by functioning as activin A (ActA). Mechanistically, ActA induces bone morphogenetic protein receptor type 2 (BMPRII) internalization and targeting to lysosomes for degradation, resulting in BMPRII signal deficiency in ECs. When endothelial ActA-BMPRII link is overdriven in mice, hypoxia-induced pulmonary hypertension was exacerbated, whereas conditional knockout of INHBA in ECs prevents the progression of pulmonary hypertension. These data collectively indicate a critical role for the dysregulated endothelial ActA-BMPRII link in the progression of pulmonary hypertension, and thus endothelial INHBA/ActA is an attractive pharmacotherapeutic target for the treatment of PAH. For cells, commercially available human microvascular ECs of the dermis and the lungs (MVECs-D and MVECs-L), human pulmonary artery ECs (PAECs) and human coronary artery ECs (CAEC) (n=1-2) were cultured until they reach subconfluency (~90%), at which the RNA was extracted using TRIzol. For human tissue samples, RNAs for 5-donor pooled human lungs, heart, kidneys, and liver of were commercially obtained. Labeled cRNA was prepared using 1 vial of the mixed RNAs for each sample according to the One-Color Microarray-Based Gene Expression Analysis ver. 6.5 protocol, and 600 ng of cRNAs was used for hybridization with microarray chips (SurePrint G3 Human GE; G4851B) for 17 h. After washing, signals were scanned using an Agilent DNA microarray scanner, and the data were analyzed using Feature Extraction ver. 10.7.1.1 (Agilent Technologies).