DJ, Majka SM. ABCG2 pos lung mesenchymal stem cells are a novel pericyte subpopulation that contributes to fibrotic remodeling.

Pulmonary vascular remodeling and oxidative stress are common to many adult lung diseases. However, little is known about the relevance of lung mesenchymal stem cells (MSCs) in these processes. We tested the hypothesis that dysfunctional lung MSCs directly participate in remodeling of the microcirculation. We employed a genetic model to deplete extracellular superoxide dismutase (EC-SOD) in lung MSCs coupled with lineage tracing analysis. We crossed floxpsod3 and mT/mG reporter mice to a strain expressing Cre recombinase under the control of the ABCG2 promoter. We demonstrated In vivo that depletion of EC-SOD in lung MSCs resulted in their contribution to microvascular remodeling in the smooth muscle actin positive layer. We further characterized lung MSCs to be multipotent vascular precursors, capable of myofibroblast, endothelial and pericyte differentiation in vitro. EC-SOD deficiency in cultured lung MSCs accelerated proliferation and apoptosis, restricted colony-forming ability, multilineage differentiation potential and promoted the transition to a contractile phenotype. Further studies correlated cell dysfunction to alterations in canonical Wnt/β-catenin signaling, which were more evident under conditions of oxidative stress. Our data establish that lung MSCs are a multipotent vascular precursor population, a population which has the capacity to participate in vascular remodeling and their function is likely regulated in part by the Wnt/β-catenin signaling pathway. These studies highlight an important role for microenviromental regulation of multipotent MSC function as well as their potential to contribute to tissue remodeling.

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 ...

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Rationale: Idiopathic pulmonary arterial hypertension (IPAH) is usually without an identified genetic cause, despite clinical and molecular similarity to bone morphogenetic protein receptor type 2 mutation-associated heritable pulmonary arterial hypertension (PAH). There is phenotypic heterogeneity in IPAH, with a minority of patients showing long-term improvement with calcium channel-blocker therapy. Objectives: We sought to identify gene variants (GVs) underlying IPAH and determine whether GVs differ in vasodilator-responsive IPAH (VR-PAH) versus vasodilator-nonresponsive IPAH (VN-PAH).Methods: We performed whole-exome sequencing (WES) on 36 patients with IPAH: 17 with VR-PAH and 19 with VN-PAH. Wnt pathway differences were explored in human lung fibroblasts.Measurements and Main Results: We identified 1,369 genes with 1,580 variants unique to IPAH. We used a gene ontology approach to analyze variants and identified overrepresentation of several pathways, including cytoskeletal function and ion binding. By mapping WES data to prior genome-wide association study data, Wnt pathway genes were highlighted. Using the connectivity map to define genetic differences between VR-PAH and VN-PAH, we found enrichment in vascular smooth muscle cell contraction pathways and greater genetic variation in VR-PAH versus VN-PAH. Using human lung fibroblasts, we found increased stimulated Wnt activity in IPAH versus controls.Conclusions: A pathway-based analysis of WES data in IPAH demonstrated multiple rare GVs that converge on key biological pathways, such as cytoskeletal function and Wnt signaling pathway. Vascular smooth muscle contraction-related genes were enriched in VR-PAH, suggesting a potentially different genetic predisposition for VR-PAH. This pathway-based approach may be applied to next-generation sequencing data in other diseases to uncover the contribution of unexpected or multiple GVs to a phenotype.