Identification of a common Wnt-associated genetic signature across multiple cell types in pulmonary arterial hypertension

Abstract

May 28, 2014; doi:10.1152/ajpcell.00057.2014.-Understanding differences in gene expression that increase risk for pulmonary arterial hypertension (PAH) is essential to understanding the molecular basis for disease. Previous studies on patient samples were limited by end-stage disease effects or by use of nonadherent cells, which are not ideal to model vascular cells in vivo. These studies addressed the hypothesis that pathological processes associated with PAH may be identified via a genetic signature common across multiple cell types. Expression array experiments were initially conducted to analyze cell types at different stages of vascular differentiation (mesenchymal stromal and endothelial) derived from PAH patient-specific induced pluripotent stem (iPS) cells. Molecular pathways that were altered in the PAH cell lines were then compared with those in fibroblasts from 21 patients, including those with idiopathic and heritable PAH. Wnt was identified as a target pathway and was validated in vitro using primary patient mesenchymal and endothelial cells. Taken together, our data suggest that the molecular lesions that cause PAH are present in all cell types evaluated, regardless of origin, and that stimulation of the Wnt signaling pathway was a common molecular defect in both heritable and idiopathic PAH. pulmonary arterial hypertension; gene array; induced pluripotent stem cell; mesenchymal stromal cell; endothelial cell; heritable pulmonary arterial hypertension; idiopathic pulmonary arterial hypertension; Wnt signaling PULMONARY ARTERIAL HYPERTENSION (PAH) is characterized by vascular remodeling, including endothelial cell (EC) dysfunction and occlusion or rarefaction of the peripheral pulmonary microvasculature. More recently, the contribution of multipotent mesenchymal stromal cells (MSC) to muscularization of microvessels has been described (13). The interactions between the lung microenvironment, vascular EC, and MSC during remodeling in PAH remain unclear. All forms of PAH have a high mortality rate, despite current therapeutic options.Deregulated bone morphogenetic protein (BMP) receptor type II (BMPR2) signaling is strongly associated with the development of PAH in both heritable (BMPR2 mut and Cav1 mut ) and idiopathic cases, although the molecular mechanisms through which BMPR2 derangement promotes PAH are unknown. Unfortunately, most rodent models of PAH do not precisely recapitulate the disease pathology; these models display less substantial pulmonary vascular remodeling in both proximal arteries and distal microvasculature, significantly slowing drug discovery efforts. Current in vitro models overexpressing mutant BMPR2 in cell types of interest are complicated by persistent retention of wild-type (WT) signaling. Moreover, human PAH tissue is limited in quantity, and specimens are typically obtained posttransplant or at autopsy, which limits conclusions about disease initiation and propagation. Previous global gene expression analyses using patient samples to identify risk factors for PAH have ...

Disruption of lineage specification in adult pulmonary mesenchymal progenitor cells promotes microvascular dysfunction

“…Our current and previous studies directly link increased β-catenin signaling with deregulated BMPR signaling (22,79). These data suggest that intact BMP signaling promotes the differentiation of pulmonary mesenchymal pericyte progenitors and suppresses Wnt/β-catenin signaling, which is required for the selfrenewal and proliferation of progenitor pools.…”

“…The cells were allowed to remain in 20% serum medium for 24 hours. After 24 hours, the medium was changed to 20% serum treatment medium containing DM (5 μM), DMH1 (10 μM), or DMSO vehicle (22). RNA lysates were collected at 48 hours and protein lysates…”

“…Reactivation of the developmental signaling pathways is often associated with abnormal lineage specification and disease pathology in adult tissues. Coordinated Wnt and BMPR2 signaling influence the proliferation and survival of endothelium and smooth muscle during angiogenesis as well as vascular remodeling of the pulmonary circulation (20)(21)(22). We therefore sought to determine how their coordinated signaling in resident lung ABCG2 + mesenchymal precursors regulates pulmonary microvascular homeostasis in vivo and in vitro.…”

Critical Genomic Networks and Vasoreactive Variants in Idiopathic Pulmonary Arterial Hypertension

“…Rationale for selection of the variants and their frequency is in the online supplement (Table E6). We further sought to determine if any of the variant pathways identified through WES have functional consequences using skin fibroblasts cultured from patients with IPAH (32). Gene ontology analysis was performed via Webgestalt (36) using genes with altered regulation in skin fibroblasts in patients with IPAH compared with control subjects.…”

Putting skin in the game: dermis-derived stem cells provide insight into familial pulmonary hypertension

“…The numbers of subjects within groups for individual experiments were small, with no data given on the severity of PAH or on comorbidities that may have affected the condition of skin fibroblasts. It is unclear if they evaluated for abnormalities of wnt signaling in iPSC-EC that others have reported to exist in FPAH (9). A parallel study arm where iPSC are differentiated into vascular smooth muscle cells to address other key events of PAH pathogenesis would also have been of interest.…”