RNA Sequencing Analysis Detection of a Novel Pathway of Endothelial Dysfunction in Pulmonary Arterial Hypertension

Objective.Pulmonary endothelial dysfunction and increased reflection of pulmonary pressure waves have been reported in pulmonary arterial hypertension (PAH). However, the systemic vascular involvement is not fully understood. Our study focused on the systemic arterial stiffness and endothelial involvement in idiopathic and scleroderma associated PAH.Methods.Peripheral arterial stiffness and endothelial function were evaluated in 38 patients with idiopathic (n = 28) and scleroderma associated (n = 10) PAH, and 21 control subjects (13 healthy; 8 with scleroderma and normal pulmonary pressure). All participants underwent clinical and cardiopulmonary evaluation. Arterial stiffness was measured through the fingertip tonometry derived augmentation index (AI), which is the boost increase in the late systolic pressure wave after the initial systolic shoulder. Endothelial function was measured by forearm blood flow dilatation response to brachial artery occlusion by a noninvasive plethysmograph (EndoPAT 2000), which is associated with nitric oxide-dependent vasodilatation and yields a peripheral arterial tone (PAT) ratio.Results.Mean systolic pulmonary pressure was 70.5 ± 21.6 mm Hg (idiopathic-PAH) and 69.3 ± 20 mm Hg (scleroderma-PAH). AI was higher in scleroderma patients (10.5% ± 19.6% in healthy controls, 9.0% ± 21.5% in idiopathic-PAH, 20.1% ± 19.1% in scleroderma-PAH, and 24.4% ± 18.9% in scleroderma-controls; nonsignificant). PAT ratio was significantly lower (p < 0.05) than control values in idiopathic-PAH and scleroderma-PAH (PAT ratio: control 2.20 ± 0.25; idiopathic 1.84 ± 0.51; scleroderma 1.66 ± 0.66). AI was not correlated to endothelial dysfunction. There were no differences between the 2 PAH patient groups in age, body mass index, New York Heart Association classification, or 6-min walk test.Conclusion.Our study shows a trend towards increased arterial stiffness in scleroderma (nonsignificant), and also peripheral endothelial dysfunction in idiopathic-PAH and in scleroderma-PAH. These findings suggest involvement of different vessels in scleroderma-PAH compared to idiopathic-PAH.

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Peripheral endothelial dysfunction in patients with pulmonary arterial hypertension

Respiratory Medicine ◽

10.1016/j.rmed.2008.06.014 ◽

2008 ◽

Vol 102 (12) ◽

pp. 1791-1796 ◽

Cited By ~ 35

Author(s):

Nir Peled ◽

Daniele Bendayan ◽

David Shitrit ◽

Ben Fox ◽

Liora Yehoshua ◽

...

Keyword(s):

Pulmonary Arterial Hypertension ◽

Endothelial Dysfunction ◽

Arterial Hypertension ◽

Pulmonary Arterial

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The FGL2 prothrombinase contributes to the pathological process of experimental pulmonary hypertension

Journal of Applied Physiology ◽

10.1152/japplphysiol.00396.2019 ◽

2019 ◽

Vol 127 (6) ◽

pp. 1677-1687

Author(s):

Cheng Fan ◽

Jue Wang ◽

Chaoqin Mao ◽

Wenzhu Li ◽

Kun Liu ◽

...

Keyword(s):

Pulmonary Hypertension ◽

Pulmonary Arterial Hypertension ◽

Endothelial Dysfunction ◽

Arterial Hypertension ◽

Vascular Remodeling ◽

Thrombus Formation ◽

Idiopathic Pulmonary Arterial Hypertension ◽

Potential Therapeutic Target ◽

Pulmonary Arterial

In situ thrombus formation is one of the major pathological features of pulmonary hypertension (PH). The mechanism of in situ thrombosis has not been clearly identified. Fibrinogen-like protein 2 (FGL2) prothrombinase is an immune coagulant that can cleave prothrombin to thrombin, which then converts fibrinogen into fibrin. This mechanism triggers in situ thrombus formation directly, bypassing both the intrinsic and extrinsic coagulation pathways. FGL2 prothrombinase is mainly expressed in endothelial cells and mediates multiple pathological processes. This implies that it may also play a role in PH. In this study, we examined the expression of FGL2 in idiopathic pulmonary arterial hypertension (IPAH) patients, and in monocrotaline-induced rat and hypoxia-induced mouse PH models. Fgl2−/− mice were used to evaluate the development of PH and explore associated pathological changes. These included in situ thrombosis, vascular remodeling, and endothelial apoptosis. Following these analyses, we examined possible signaling pathways downstream of FGL2 in PH. We show FGL2 is upregulated in pulmonary vascular endothelium in human IPAH and in two animal PH models. Genetic knockout of Fgl2 limited the development of PH, indicated by decreased in situ thrombus formation, less vascular remodeling, and reduced endothelial dysfunction. In addition, loss of FGL2 downregulated PAR1 (proteinase-activated receptor 1) expression and decreased the overactivation and consumption of platelets in hypoxia-induced PH. These results indicate FGL2 participate in the development of PH and loss of FGL2 could attenuate PH by reducing in situ thrombosis and suppressing PAR1 signaling. Thus we provide evidence that suggests FGL2 prothrombinase presents a potential therapeutic target for clinical treatment of PH. NEW & NOTEWORTHY This is the first study to demonstrate that fibrinogen-like protein 2 (FGL2) participates in the pathological progression of pulmonary hypertension (PH) in human idiopathic pulmonary arterial hypertension, a monocrotaline rat PH model, and a hypoxia mouse PH model. Genetic knockout of Fgl2 significantly limited the development of PH indicated by reduced in situ thrombosis, vascular remodeling, and endothelial dysfunction, and suppressed PAR1 (proteinase-activated receptor 1) signaling and overactivation of platelets on PH. These results suggest FGL2 presents a potential therapeutic target for clinical treatment of PH.

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Faculty Opinions recommendation of Nitric oxide deficiency and endothelial dysfunction in pulmonary arterial hypertension.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718026516.793479771 ◽

2013 ◽

Author(s):

Vinicio de Jesus Perez

Keyword(s):

Nitric Oxide ◽

Pulmonary Arterial Hypertension ◽

Endothelial Dysfunction ◽

Arterial Hypertension ◽

Nitric Oxide Deficiency ◽

Pulmonary Arterial

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Abstract 14257: Endothelial SOX17 Deficiency Induces Pulmonary Arterial Hypertension

Circulation ◽

10.1161/circ.142.suppl_3.14257 ◽

2020 ◽

Vol 142 (Suppl_3) ◽

Author(s):

Zhiyu Dai ◽

Dan Yi ◽

BIN LIU ◽

Shuai Li

Keyword(s):

Cell Cycle ◽

Cell Proliferation ◽

Pulmonary Arterial Hypertension ◽

Arterial Hypertension ◽

Vascular Remodeling ◽

Molecular Mechanisms ◽

Association Studies ◽

Genome Wide Association Studies ◽

Sequencing Analysis ◽

Pulmonary Arterial

Introduction: Pulmonary arterial hypertension (PAH) is a disaster disease characterized by obliterative vascular remodeling and persistent increase of vascular resistance, leading to right heart failure and premature death. Understanding the cellular and molecular mechanisms will help develop novel therapeutic approaches for PAH patients. Hypothesis: Human genome-wide association studies identified that SOX17 locus variants are associated with PAH. SOX17 mutation is also found in patients with PAH. We hypothesis that endothelial SOX17 deficiency contributes to the pathogenesis of PAH. Methods: Mice with EndoSCL-CreERT mediated deletion of Sox17 ( Sox17 iCKO ) were generated. Sox17 iCKO and Sox17 f/f mice after tamoxifen injection were incubated with hypoxia (10% O 2 ) for 3 weeks to induced PAH. Hemodynamics and histological examination were measured to determine the PAH phenotypes and vascular remodeling. EC proliferation and apoptosis were assessed in SiRNA-mediated SOX17 knockdown in human lung microvascular endothelial cells (hLMVECs). The RNA-sequencing analysis was performed to understand the molecular mechanisms of SOX17 deficiency in ECs. Results: Sox17 iCKO mice exhibited exaggerative PAH evident by the increase of RVSP and RV hypertrophy after hypoxia treatment compared to Sox17 f/f WT mice. SOX17 knockdown in hLMVECs induced cell proliferation and reduced starvation-induced apoptosis. RNA-seq analysis and DAVID pathway analysis demonstrated that there was dysregulation of cell proliferation-related genes, which are enriched in the pathways related to cell cycle, cell division, and mitotic cell cycle. Transcriptional factor, target, and motif discovery analysis of the dysregulated gene set revealed the involvement of transcriptional factors FOXM1 and E2F1. siRNA mediated knockdown of E2F1 but not FOXM1 normalized SOX17 deficiency-induced hLMVECs proliferation and anti-apoptosis. Conclusions: Our study demonstrated that endothelial SOX17 deficiency exaggerates hypoxia-induced PAH. Loss of SOX17 promotes EC proliferation and anti-apoptosis via the upregulation of transcription factor E2F1.

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Abstract 15544: Implication of the Histone Methyltransferase “G9a” in Pulmonary Arterial Hypertension

Circulation ◽

10.1161/circ.142.suppl_3.15544 ◽

2020 ◽

Vol 142 (Suppl_3) ◽

Author(s):

Charifa awada ◽

Karima Habbout ◽

Valerie Nadeau ◽

Sandra Breuils-Bonnet ◽

Roxane Paulin ◽

...

Keyword(s):

Pulmonary Arterial Hypertension ◽

Arterial Hypertension ◽

Cholesterol Metabolism ◽

Cholesterol Biosynthesis ◽

Histone Methyltransferase ◽

Sequencing Analysis ◽

Apoptosis Resistance ◽

Cancer Pathogenesis ◽

Therapeutic Benefits ◽

Pulmonary Arterial

Rationale: Pulmonary arterial hypertension (PAH) is a cardiopulmonary disorder characterized by elevation of pulmonary arterial (PA) pressure and premature death. PA smooth muscle cells (PASMCs) from PAH patients present a cancer-like hyperproliferative and apoptosis-resistant phenotype contributing to remodeling of distal PAs. Although epigenetic alterations contribute to PAH development, one important challenge is defining which genes are the drivers . A growing body of literature points to the role of an epigenetic factor called G9a in cancer pathogenesis. Indeed, G9a is a histone methyltransferase overexpressed in many cancers promoting cell proliferation and survival. Given the similarities between PAH and cancer, it is of interest to determine whether G9a is implicated in PAH. We thus hypothesized that G9a inhibition reduces the pro-proliferative and apoptosis resistance phenotype of PAH-PASMCs. Methods and Results: Using Western blot (WB) and immunofluorescence (IF), we showed that G9a is overexpressed in distal PAs and isolated PASMCs from PAH patients (n= 6-14, p<0.01). Similarly, G9a was increased (WB and IF, p<0.05) in two models mimicking the disease; namely the monocrotaline rat and mice exposed to chronic hypoxia. In vitro, we found that pharmacological inhibition of G9a using BIX01294 and UNC0642 reduces PAH-PASMC proliferation (Ki67 and EdU assays, p<0.001) and survival (Annexin V assay p<0.001). Through RNA sequencing analysis conducted in PAH-PASMCs treated or not with BIX01294, we found that upregulated differentially expressed genes (DEGs) were enriched in cholesterol biosynthesis, autophagy-lysosome and ER stress-induced apoptotic pathways. However, downregulated DEGs were involved in cell cycle and fibrosis-related processes. Consistently, inhibition of G9a generates numerous cytoplasmic vacuoles positive for LC3-II and p62 (WB, IF), thus suggesting that the inhibition of G9a induces cell death by altering cholesterol metabolism-dependent autophagy. Conclusion: We showed for the first time that G9a is overexpressed in PAH contributing to the pro-proliferative and anti-apoptotic phenotype of PAH-PASMCs. Current experiments aim to determine whether G9a inhibition provides therapeutic benefits in PAH.

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Endothelial dysfunction in pulmonary arterial hypertension: an evolving landscape (2017 Grover Conference Series)

Pulmonary Circulation ◽

10.1177/2045893217752912 ◽

2017 ◽

Vol 8 (1) ◽

pp. 204589321775291 ◽

Cited By ~ 36

Author(s):

Benoît Ranchoux ◽

Lloyd D. Harvey ◽

Ramon J. Ayon ◽

Aleksandra Babicheva ◽

Sebastien Bonnet ◽

...

Keyword(s):

Pulmonary Arterial Hypertension ◽

Endothelial Dysfunction ◽

Arterial Hypertension ◽

Molecular Mechanisms ◽

Right Heart Failure ◽

Vascular Pathology ◽

Mesenchymal Transition ◽

Conference Series ◽

Major Player ◽

Pulmonary Arterial

Endothelial dysfunction is a major player in the development and progression of vascular pathology in pulmonary arterial hypertension (PAH), a disease associated with small vessel loss and obstructive vasculopathy that leads to increased pulmonary vascular resistance, subsequent right heart failure, and premature death. Over the past ten years, there has been tremendous progress in our understanding of pulmonary endothelial biology as it pertains to the genetic and molecular mechanisms that orchestrate the endothelial response to direct or indirect injury, and how their dysregulation can contribute to the pathogenesis of PAH. As one of the major topics included in the 2017 Grover Conference Series, discussion centered on recent developments in four areas of pulmonary endothelial biology: (1) angiogenesis; (2) endothelial-mesenchymal transition (EndMT); (3) epigenetics; and (4) biology of voltage-gated ion channels. The present review will summarize the content of these discussions and provide a perspective on the most promising aspects of endothelial dysfunction that may be amenable for therapeutic development.

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Pulmonary arterial hypertension in systemic sclerosis: Diagnosis and treatment according to the European Society of Cardiology and European Respiratory Society 2015 guidelines

Journal of Scleroderma and Related Disorders ◽

10.1177/2397198318808998 ◽

2018 ◽

Vol 4 (1) ◽

pp. 35-42 ◽

Cited By ~ 4

Author(s):

Nicola Giordano ◽

Claudio Corallo ◽

Chiara Chirico ◽

Angelica Brazzi ◽

Adriana Marinetti ◽

...

Keyword(s):

Pulmonary Hypertension ◽

Systemic Sclerosis ◽

Pulmonary Arterial Hypertension ◽

Endothelial Dysfunction ◽

Arterial Hypertension ◽

Correct Diagnosis ◽

European Respiratory Society ◽

Long Term Outcome ◽

Screening Programs ◽

Pulmonary Arterial

Scleroderma (systemic sclerosis) is an autoimmune connective tissue disease which presents endothelial dysfunction and fibroblast dysregulation, resulting in vascular and fibrotic disorders. Pulmonary hypertension is frequent in patients with systemic sclerosis: the natural evolution of the disease can induce the development of different forms of pulmonary hypertension, representing one of the main causes of death. Among the different forms of pulmonary hypertension in systemic sclerosis, pulmonary arterial hypertension is the most frequent one (rate of occurrence is estimated between 7% and 12%). This pulmonary vascular complication should be treated with a combination of drugs that is able to counteract endothelial dysfunction, antagonizing the endothelin-1 system and replacing prostaglandin I2 and nitric oxide activity. A correct diagnosis is mandatory, because it is possible only for pulmonary arterial hypertension to use specific drugs that are able to control the symptomatic condition and the evolution of the disease. According to the most recent guidelines, for the patients with systemic sclerosis, also without pulmonary hypertension symptoms, echocardiography screening for the detection of pulmonary hypertension is recommended. Pulmonary arterial hypertension screening programs in systemic sclerosis patients is able to identify milder forms of the disease, allowing earlier management and better long-term outcome.

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