MEF2C-MYOCD and Leiomodin1 Suppression by miRNA-214 Promotes Smooth Muscle Cell Phenotype Switching in Pulmonary Arterial Hypertension.

BACKGROUND: Vascular hyperproliferative disorders are characterized by excessive smooth muscle cell (SMC) proliferation leading to vessel remodeling and occlusion. In pulmonary arterial hypertension (PAH), SMC phenotype switching from a terminally differentiated contractile to synthetic state is gaining traction as our understanding of the disease progression improves. While maintenance of SMC contractile phenotype is reportedly orchestrated by a MEF2C-myocardin (MYOCD) interplay, little is known regarding molecular control at this nexus. Moreover, the burgeoning interest in microRNAs (miRs) provides the basis for exploring their modulation of MEF2C-MYOCD signaling, and in turn, a pro-proliferative, synthetic SMC phenotype. We hypothesized that suppression of SMC contractile phenotype in pulmonary hypertension is mediated by miR-214 via repression of the MEF2C-MYOCD-leiomodin1 (LMOD1) signaling axis. METHODS AND RESULTS: In SMCs isolated from a PAH patient cohort and commercially obtained hPASMCs exposed to hypoxia, miR-214 expression was monitored by qRT-PCR. miR-214 was upregulated in PAH- vs. control subject hPASMCs as well as in commercially obtained hPASMCs exposed to hypoxia. These increases in miR-214 were paralleled by MEF2C, MYOCD and SMC contractile protein downregulation. Of these, LMOD1 and MEF2C were directly targeted by the miR. Mir-214 overexpression mimicked the PAH profile, downregulating MEF2C and LMOD1. AntagomiR-214 abrogated hypoxia-induced suppression of the contractile phenotype and its attendant proliferation. Anti-miR-214 also restored PAH-PASMCs to a contractile phenotype seen during vascular homeostasis. CONCLUSIONS: Our findings illustrate a key role for miR-214 in modulation of MEF2C-MYOCD-LMOD1 signaling and suggest that an antagonist of miR-214 could mitigate SMC phenotype changes and proliferation in vascular hyperproliferative disorders including PAH

The role of nicotinamide adenine dinucleotide phosphate (reduced form) oxidase in endothelial activation in sepsis /

Septic shock is a leading cause of death in intensive care units. As part of the septic process, the endothelium becomes activated and propagates the septic condition. It has become evident that reactive oxygen species (ROS) are involved in the signaling of mediators of sepsis, such as tumor necrosis factor-alpha (TNF-alpha) and the lipopolysaccharide coating of gram-negative bacteria (LPS). An important source of these ROS is NADPH oxidase, which is a ubiquitously expressed enzyme complex that also exists in endothelial cells (EC). We showed that O2- from NADPH oxidase was important for LPS, as well as TNF-alpha, induction of two markers of an activated endothelium, interleukin-8 (IL-8) and intercellular adhesion molecule-1 (ICAM-1) in human umbilical vein endothelial cells (HUVEC).Expression of a gene can be increased by a rise in transcription as well as post-transcriptional changes, such as mRNA stability modifications. We assessed the role of NADPH oxidase in this process and found a complex interaction. Although LPS increases IL-8 transcription, it also destabilizes IL-8 mRNA in a p38 and extracellular signal-regulated kinase (ERK) MAPK dependent manner, which was only evident after blocking NADPH oxidase. This regulation involved the mRNA de-stabilizing factor tristetraprolin (TTP). In contrast, TNF-alpha enhanced the stability of IL-8, IL-6 and ICAM-1 mRNA in a p38 MAPK dependent, but NADPH oxidase independent manner. Furthermore, LPS did not have an effect on mRNA stability of IL-6 or ICAM-1 in our system. Thus, we conclude from our studies that the NAPDH oxidase is important for the induction of inflammatory molecules in LPS and TNF-alpha treated EC and is also involved in mRNA stability regulation of these molecules in a signal and gene specific fashion

Recent advancements in pulmonary arterial hypertension and right heart failure research: overview of selected abstracts from ATS2020 and emerging COVID-19 research

Each year the American Thoracic Society (ATS) Conference brings together scientists who conduct basic, translational and clinical research to present on the recent advances in the field of respirology. Due to the Coronavirus Disease of 2019 (COVID-19) pandemic, the ATS2020 Conference was held online in a series of virtual meetings. In this review, we focus on the breakthroughs in pulmonary hypertension research. We have selected 11 of the best basic science abstracts which were presented at the ATS2020 Assembly on Pulmonary Circulation mini-symposium “What’s New in Pulmonary Arterial Hypertension (PAH) and Right Ventricular (RV) Signaling: Lessons from the Best Abstracts,” reflecting the current state of the art and associated challenges in PH. Particular emphasis is placed on understanding the mechanisms underlying RV failure, the regulation of inflammation, and the novel therapeutic targets that emerged from preclinical research. The pathologic interactions between pulmonary hypertension, right ventricular function and COVID-19 are also discussed

Specificity Protein 1-Mediated Promotion of CXCL12 Advances Endothelial Cell Metabolism and Proliferation in Pulmonary Hypertension

Pulmonary arterial hypertension (PAH) is a rare yet devastating and incurable disease with few treatment options. The underlying mechanisms of PAH appear to involve substantial cellular proliferation and vascular remodeling, causing right ventricular overload and eventual heart failure. Recent evidence suggests a significant seminal role of the pulmonary endothelium in the initiation and promotion of PAH. Our previous work identified elevated reactive oxygen species (ROS)-producing enzyme NADPH oxidase 1 (NOX1) in human pulmonary artery endothelial cells (HPAECs) of PAH patients promoting endothelial cell proliferation in vitro. In this study, we interrogated chemokine CXCL12′s (aka SDF-1) role in EC proliferation under the control of NOX1 and specificity protein 1 (Sp1). We report here that NOX1 can drive hypoxia-induced endothelial CXCL12 expression via the transcription factor Sp1 leading to HPAEC proliferation and migration. Indeed, NOX1 drove hypoxia-induced Sp1 activation, along with an increased capacity of Sp1 to bind cognate promoter regions in the CXCL12 promoter. Sp1 activation induced elevated expression of CXCL12 in hypoxic HPAECs, supporting downstream induction of expression at the CXCL12 promoter via NOX1 activity. Pathological levels of CXCL12 mimicking those reported in human PAH patient serum restored EC proliferation impeded by specific NOX1 inhibitor. The translational relevance of our findings is highlighted by elevated NOX1 activity, Sp1 activation, and CXCL12 expression in explanted lung samples from PAH patients compared to non-PAH controls. Analysis of phosphofructokinase, glucose-6-phosphate dehydrogenase, and glutaminase activity revealed that CXCL12 induces glutamine and glucose metabolism, which are foundational to EC cell proliferation. Indeed, in explanted human PAH lungs, demonstrably higher glutaminase activity was detected compared to healthy controls. Finally, infusion of recombinant CXCL12 into healthy mice amplified pulmonary arterial pressure, right ventricle remodeling, and elevated glucose and glutamine metabolism. Together these data suggest a central role for a novel NOX1-Sp1-CXCL12 pathway in mediating PAH phenotype in the lung endothelium

Persistence of pulmonary hypertension in patients undergoing ventricular assist devices and orthotopic heart transplantation

Abstract Pulmonary hypertension (PH) is common in advanced heart failure and often improves quickly after left ventricular assist device (VAD) implantation or orthotopic heart transplantation (OHT), but long‐term effects and outcomes are not well‐described. This study evaluated PH persistence after VAD as destination therapy (VAD‐DT), bridge to transplant (VAD‐OHT), or OHT‐alone. The study constituted a retrospective review of patients who underwent VAD‐DT (n = 164), VAD‐OHT (n = 111), or OHT‐alone (n = 138) at a single tertiary‐care center. Right heart catheterization (RHC) data was collected pre‐, post‐intervention (VAD and/or OHT), and 1‐year from final intervention (latest‐RHC) to evaluate the longitudinal hemodynamic course of right ventricular function and pulmonary vasculature. PH (Group II and Group I) definitions were adapted from expert guidelines. All groups showed significant improvements in mean pulmonary artery pressure (mPAP), pulmonary artery wedge pressure (PAWP), cardiac output, and pulmonary vascular resistance (PVR) at each RHC with greatest improvement at post‐intervention RHC (post‐VAD or post‐OHT). PH was reduced from 98% to 26% in VAD‐OHT, 92%−49% in VAD‐DT, and 76%−28% in OHT‐alone from preintervention to latest‐RHC. At latest‐RHC mPAP remained elevated in all groups despite normalization of PAWP and PVR. VAD‐supported patients exhibited suppressed pulmonary artery pulsatility index (PaPi < 3.7) with improvement only posttransplant at latest‐RHC. Posttransplant patients with PH at latest‐RHC (n = 60) exhibited lower survival (HR: 2.1 [95% CI: 1.3−3.4], p < 0.001). Despite an overall significant improvement in pulmonary pressures and PH proportion, a notable subset of patients exhibited PH post‐intervention. Post‐intervention PH was associated with lower posttransplant survival