The diverging roles of insulin-like growth factor binding proteins in pulmonary arterial hypertension.

Pulmonary hypertension (PH) is a progressive, severe and to date not curable disease of the pulmonary vasculature. Alterations of the insulin-like growth factor 1 (IGF-1) system are known to play a role in vascular pathologies and IGF-binding proteins (IGFBPs) are important regulators of the bioavailability and function of IGFs. In this study, we show that circulating plasma levels of IGFBP-1, IGFBP-2 and IGFBP-3 are increased in idiopathic pulmonary arterial hypertension (IPAH) patients compared to healthy individuals. These binding proteins inhibit the IGF-1 induced IGF-1 receptor (IGF1R) phosphorylation and exhibit diverging effects on the IGF-1 induced signaling pathways in human pulmonary arterial cells (i.e. healthy as well as IPAH-hPASMCs, and healthy hPAECs). Furthermore, IGFBPs are differentially expressed in an experimental mouse model of PH. In hypoxic mouse lungs, IGFBP-1 mRNA expression is decreased whereas the mRNA for IGFBP-2 is increased. In contrast to IGFBP-1, IGFBP-2 shows vaso-constrictive properties in the murine pulmonary vasculature. Our analyses show that IGFBP-1 and IGFBP-2 exhibit diverging effects on IGF-1 signaling and display a unique IGF1R-independent kinase activation pattern in human pulmonary arterial smooth muscle cells (hPASMCs), which represent a major contributor of PAH pathobiology. Furthermore, we could show that IGFBP-2, in contrast to IGFBP-1, induces epidermal growth factor receptor (EGFR) signaling, Stat-3 activation and expression of Stat-3 target genes. Based on our results, we conclude that the IGFBP family, especially IGFBP-1, IGFBP-2 and IGFBP-3, are deregulated in PAH, that they affect IGF signaling and thereby regulate the cellular phenotype in PH.

A Novel Rat Model of Mild Pulmonary Hypertension Associated with Pulmonary Venous Congestion Induced by Left Pulmonary Vein Banding.

Pulmonary hypertension (PH) associated with left heart disease (PH-LHD) is the most common form of PH. In PH-LHD, changes in the pulmonary vasculature are assumed to be mainly caused by pulmonary venous congestion. However, the underlying mechanisms of this form of PH are poorly understood. We aimed to establish a model of PH associated with pulmonary venous congestion. Wistar-Kyoto rats underwent partial occlusion of the left pulmonary vein to induce pulmonary venous congestion or sham surgery and were assessed at various time points post-surgery (3, 6, 9, 12 weeks). In vivo cardiopulmonary phenotyping was performed by using echocardiography along with heart catheterization. Histomorphometry methods were used to assess pulmonary vascular remodeling (e.g., wall thickness, degree of muscularization). Left pulmonary vein banding (PVB) resulted in mildly elevated right ventricular systolic pressure and moderate right ventricular hypertrophy. In PVB rats, small- and medium-sized pulmonary vessels in the left lung were characterized by increased wall thickness and muscularization. Taken together, our data demonstrate that left PVB-induced pulmonary venous congestion is associated with pulmonary vascular remodeling and mild PH.

Mast Cells in Cardiac Remodeling: Focus on the Right Ventricle.

In response to various stressors, cardiac chambers undergo structural remodeling. Long-term exposure of the right ventricle (RV) to pressure or volume overload leads to its maladaptive remodeling, associated with RV failure and increased mortality. While left ventricular adverse remodeling is well understood and therapeutic options are available or emerging, RV remodeling remains underexplored, and no specific therapies are currently available. Accumulating evidence implicates the role of mast cells in RV remodeling. Mast cells produce and release numerous inflammatory mediators, growth factors and proteases that can adversely affect cardiac cells, thus contributing to cardiac remodeling. Recent experimental findings suggest that mast cells might represent a potential therapeutic target. This review examines the role of mast cells in cardiac remodeling, with a specific focus on RV remodeling, and explores the potential efficacy of therapeutic interventions targeting mast cells to mitigate adverse RV remodeling.

Alveolar macrophage-expressed Plet1 is a driver of lung epithelial repair after viral pneumonia.

Influenza A virus (IAV) infection mobilizes bone marrow-derived macrophages (BMDM) that gradually undergo transition to tissue-resident alveolar macrophages (TR-AM) in the inflamed lung. Combining high-dimensional single-cell transcriptomics with complex lung organoid modeling, in vivo adoptive cell transfer, and BMDM-specific gene targeting, we found that transitioning ("regenerative") BMDM and TR-AM highly express Placenta-expressed transcript 1 (Plet1). We reveal that Plet1 is released from alveolar macrophages, and acts as important mediator of macrophage-epithelial cross-talk during lung repair by inducing proliferation of alveolar epithelial cells and re-sealing of the epithelial barrier. Intratracheal administration of recombinant Plet1 early in the disease course attenuated viral lung injury and rescued mice from otherwise fatal disease, highlighting its therapeutic potential.

Surgical protocol for pulmonary artery banding in mice to generate a model of pressure-overload-induced right ventricular failure.

Right ventricular failure (RVF) is the leading cause of death in patients with pulmonary hypertension. Here, we present a protocol for pulmonary artery banding in mice to generate a model of pressure-overload-induced RVF. We describe steps for anesthesia of mice, endotracheal intubation, and pulmonary artery banding surgery. We then detail procedures for phenotyping and analysis. Our approach does not involve complete blockage of the pulmonary flow during clip placement and is, therefore, associated with low intraoperative mortality. For complete details on the use and execution of this protocol, please refer to Veith et al. (2020).1.

Pharmacology and Rationale for Seralutinib in the Treatment of Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is a complex disorder characterized by vascular remodeling and a consequent increase in pulmonary vascular resistance. The histologic hallmarks of PAH include plexiform and neointimal lesions of the pulmonary arterioles, which are composed of dysregulated, apoptosis-resistant endothelial cells and myofibroblasts. Platelet-derived growth factor receptors (PDGFR) α and ß, colony stimulating factor 1 receptor (CSF1R), and mast/stem cell growth factor receptor kit (c-KIT) are closely related kinases that have been implicated in PAH progression. In addition, emerging data indicate significant crosstalk between PDGF signaling and the bone morphogenetic protein receptor type 2 (BMPR2)/transforming growth factor ß (TGFß) receptor axis. This review will discuss the importance of the PDGFR-CSF1R-c-KIT signaling network in PAH pathogenesis, present evidence that the inhibition of all three nodes in this kinase network is a potential therapeutic approach for PAH, and highlight the therapeutic potential of seralutinib, currently in development for PAH, which targets these pathways.

Osteopontin in Pulmonary Hypertension.

Pulmonary hypertension (PH) is a pathological condition with multifactorial etiology, which is characterized by elevated pulmonary arterial pressure and pulmonary vascular remodeling. The underlying pathogenetic mechanisms remain poorly understood. Accumulating clinical evidence suggests that circulating osteopontin may serve as a biomarker of PH progression, severity, and prognosis, as well as an indicator of maladaptive right ventricular remodeling and dysfunction. Moreover, preclinical studies in rodent models have implicated osteopontin in PH pathogenesis. Osteopontin modulates a plethora of cellular processes within the pulmonary vasculature, including cell proliferation, migration, apoptosis, extracellular matrix synthesis, and inflammation via binding to various receptors such as integrins and CD44. In this article, we provide a comprehensive overview of the current understanding of osteopontin regulation and its impact on pulmonary vascular remodeling, as well as consider research issues required for the development of therapeutics targeting osteopontin as a potential strategy for the management of PH.

Notch1 Induces Defective Epithelial Surfactant Processing and Pulmonary Fibrosis.

Rationale: Although type II alveolar epithelial cells (AEC2s) are chronically injured in idiopathic pulmonary fibrosis (IPF), they contribute to epithelial regeneration in IPF. Objectives: We hypothesized that Notch signaling may contribute to AEC2 proliferation, dedifferentiation characterized by loss of surfactant processing machinery, and lung fibrosis in IPF. Methods: We applied microarray analysis, kinome profiling, flow cytometry, immunofluorescence analysis, western blotting, quantitative PCR, and proliferation and surface activity analysis to study epithelial differentiation, proliferation, and matrix deposition in vitro (AEC2 lines, primary murine/human AEC2s), ex vivo (human IPF-derived precision-cut lung slices), and in vivo (bleomycin and pepstatin application, Notch1 [Notch receptor 1] intracellular domain overexpression). Measurements and Main Results: We document here extensive SP-B and -C (surfactant protein-B and -C) processing defects in IPF AEC2s, due to loss of Napsin A, resulting in increased intra-alveolar surface tension and alveolar collapse and induction of endoplasmic reticulum stress in AEC2s. In vivo pharmacological inhibition of Napsin A results in the development of AEC2 injury and overt lung fibrosis. We also demonstrate that Notch1 signaling is already activated early in IPF and determines AEC2 fate by inhibiting differentiation (reduced lamellar body compartment, reduced capacity to process hydrophobic SP) and by causing increased epithelial proliferation and development of lung fibrosis, putatively via altered JAK (Janus kinase)/Stat (signal transducer and activator of transcription) signaling in AEC2s. Conversely, inhibition of Notch signaling in IPF-derived precision-cut lung slices improved the surfactant processing capacity of AEC2s and reversed fibrosis. Conclusions: Notch1 is a central regulator of AEC2 fate in IPF. It induces alveolar epithelial proliferation and loss of Napsin A and of surfactant proprotein processing, and it contributes to fibroproliferation.

Sox17 Deficiency Promotes Pulmonary Arterial Hypertension via HGF/c-Met Signaling.

BACKGROUND: In large-scale genomic studies, Sox17, an endothelial-specific transcription factor, has been suggested as a putative causal gene of pulmonary arterial hypertension (PAH); however, its role and molecular mechanisms remain to be elucidated. We investigated the functional impacts and acting mechanisms of impaired Sox17 (SRY-related HMG-box17) pathway in PAH and explored its potential as a therapeutic target. METHODS: In adult mice, Sox17 deletion in pulmonary endothelial cells (ECs) induced PAH under hypoxia with high penetrance and severity, but not under normoxia. RESULTS: Key features of PAH, such as hypermuscularization, EC hyperplasia, and inflammation in lung arterioles, right ventricular hypertrophy, and elevated pulmonary arterial pressure, persisted even after long rest in normoxia. Mechanistically, transcriptomic profiling predicted that the combination of Sox17 deficiency and hypoxia activated c-Met signaling in lung ECs. HGF (hepatocyte grow factor), a ligand of c-Met, was upregulated in Sox17-deficient lung ECs. Pharmacologic inhibition of HGF/c-Met signaling attenuated and reversed the features of PAH in both preventive and therapeutic settings. Similar to findings in animal models, Sox17 levels in lung ECs were repressed in 26.7% of PAH patients (4 of 15), while those were robust in all 14 non-PAH controls. HGF levels in pulmonary arterioles were increased in 86.7% of patients with PAH (13 of 15), but none of the controls showed that pattern. CONCLUSIONS: The downregulation of Sox17 levels in pulmonary arterioles increases the susceptibility to PAH, particularly when exposed to hypoxia. Our findings suggest the reactive upregulation of HGF/c-Met signaling as a novel druggable target for PAH treatment.

Circulating Microparticles Are Differentially Increased in Lowlanders and Highlanders with High Altitude Induced Pulmonary Hypertension during the Cold Season.

The role of microparticles (MPs) and cold in high altitude pulmonary hypertension (HAPH) remains unexplored. We investigated the impact of long-term cold exposure on the pulmonary circulation in lowlanders and high-altitude natives and the role of MPs. Pulmonary hemodynamics were evaluated using Doppler echocardiography at the end of the colder and warmer seasons. We further examined the miRNA content of MPs isolated from the study participants and studied their effects on human pulmonary artery smooth muscle (hPASMCs) and endothelial cells (hPAECs). Long-term exposure to cold environment was associated with an enhanced pulmonary artery pressure in highlanders. Plasma levels of CD62E-positive and CD68-positive MPs increased in response to cold in lowlanders and HAPH highlanders. The miRNA-210 expression contained in MPs differentially changed in response to cold in lowlanders and highlanders. MPs isolated from lowlanders and highlanders increased proliferation and reduced apoptosis of hPASMCs. Further, MPs isolated from warm-exposed HAPH highlanders and cold-exposed highlanders exerted the most pronounced effects on VEGF expression in hPAECs. We demonstrated that prolonged exposure to cold is associated with elevated pulmonary artery pressures, which are most pronounced in high-altitude residents. Further, the numbers of circulating MPs are differentially increased in lowlanders and HAPH highlanders during the colder season.

HIV and Schistosoma Co-Exposure Leads to Exacerbated Pulmonary Endothelial Remodeling and Dysfunction Associated with Altered Cytokine Landscape.

HIV and Schistosoma infections have been individually associated with pulmonary vascular disease. Co-infection with these pathogens is very common in tropical areas, with an estimate of six million people co-infected worldwide. However, the effects of HIV and Schistosoma co-exposure on the pulmonary vasculature and its impact on the development of pulmonary vascular disease are largely unknown. Here, we have approached these questions by using a non-infectious animal model based on lung embolization of Schistosoma mansoni eggs in HIV-1 transgenic (HIV) mice. Schistosome-exposed HIV mice but not wild-type (Wt) counterparts showed augmented pulmonary arterial pressure associated with markedly suppressed endothelial-dependent vasodilation, increased endothelial remodeling and vessel obliterations, formation of plexiform-like lesions and a higher degree of perivascular fibrosis. In contrast, medial wall muscularization was similarly increased in both types of mice. Moreover, HIV mice displayed an impaired immune response to parasite eggs in the lung, as suggested by decreased pulmonary leukocyte infiltration, small-sized granulomas, and augmented residual egg burden. Notably, vascular changes in co-exposed mice were associated with increased expression of proinflammatory and profibrotic cytokines, including IFN-γ and IL-17A in CD4+ and γδ T cells and IL-13 in myeloid cells. Collectively, our study shows for the first time that combined pulmonary persistence of HIV proteins and Schistosoma eggs, as it may occur in co-infected people, alters the cytokine landscape and targets the vascular endothelium for aggravated pulmonary vascular pathology. Furthermore, it provides an experimental model for the understanding of pulmonary vascular disease associated with HIV and Schistosoma co-morbidity.

Comparative kinase activity profiling of pathogenic influenza A viruses reveals new anti- and pro-viral protein kinases.

Constant evolution of influenza A viruses (IAVs) leads to the occurrence of new virus strains, which can cause epidemics and occasional pandemics. Here we compared two medically relevant IAVs, namely A/Hamburg/4/09 (H1N1pdm09) of the 2009 pandemic and the highly pathogenic avian IAV human isolate A/Thailand/1(KAN-1)/2004 (H5N1), for their ability to trigger intracellular phosphorylation patterns using a highly sensitive peptide-based kinase activity profiling approach. Virus-dependent tyrosine phosphorylations of substrate peptides largely overlap between the two viruses and are also strongly overrepresented in comparison to serine/threonine peptide phosphorylations. Both viruses trigger phosphorylations with distinct kinetics by overlapping and different kinases from which many form highly interconnected networks. As approximately half of the kinases forming a signalling hub have no known function for the IAV life cycle, we interrogated selected members of this group for their ability to interfere with IAV replication. These experiments revealed negative regulation of H1N1pdm09 and H5N1 replication by NUAK [novel (nua) kinase] kinases and by redundant ephrin A (EphA) receptor tyrosine kinases.

Adenylate Kinase 4-A Key Regulator of Proliferation and Metabolic Shift in Human Pulmonary Arterial Smooth Muscle Cells via Akt and HIF-1α Signaling Pathways.

Increased proliferation of pulmonary arterial smooth muscle cells (PASMCs) in response to chronic hypoxia contributes to pulmonary vascular remodeling in pulmonary hypertension (PH). PH shares numerous similarities with cancer, including a metabolic shift towards glycolysis. In lung cancer, adenylate kinase 4 (AK4) promotes metabolic reprogramming and metastasis. Against this background, we show that AK4 regulates cell proliferation and energy metabolism of primary human PASMCs. We demonstrate that chronic hypoxia upregulates AK4 in PASMCs in a hypoxia-inducible factor-1α (HIF-1α)-dependent manner. RNA interference of AK4 decreases the viability and proliferation of PASMCs under both normoxia and chronic hypoxia. AK4 silencing in PASMCs augments mitochondrial respiration and reduces glycolytic metabolism. The observed effects are associated with reduced levels of phosphorylated protein kinase B (Akt) as well as HIF-1α, indicating the existence of an AK4-HIF-1α feedforward loop in hypoxic PASMCs. Finally, we show that AK4 levels are elevated in pulmonary vessels from patients with idiopathic pulmonary arterial hypertension (IPAH), and AK4 silencing decreases glycolytic metabolism of IPAH-PASMCs. We conclude that AK4 is a new metabolic regulator in PASMCs interacting with HIF-1α and Akt signaling pathways to drive the pro-proliferative and glycolytic phenotype of PH.

Novel Therapeutic Targets for the Treatment of Right Ventricular Remodeling: Insights from the Pulmonary Artery Banding Model.

Right ventricular (RV) function is the main determinant of the outcome of patients with pulmonary hypertension (PH). RV dysfunction develops gradually and worsens progressively over the course of PH, resulting in RV failure and premature death. Currently, approved therapies for the treatment of left ventricular failure are not established for the RV. Furthermore, the direct effects of specific vasoactive drugs for treatment of pulmonary arterial hypertension (PAH, Group 1 of PH) on RV are not fully investigated. Pulmonary artery banding (PAB) allows to study the pathogenesis of RV failure solely, thereby testing potential therapies independently of pulmonary vascular changes. This review aims to discuss recent studies of the mechanisms of RV remodeling and RV-directed therapies based on the PAB model.

Beneficial effects of inhaled surfactant in patients with COVID-19-associated acute respiratory distress syndrome.

BACKGROUND: We have investigated the use of nebulized surfactant as a potential therapeutic option for the patients with coronavirus disease 2019 (COVID-19)-associated acute respiratory distress syndrome (ARDS) undergoing non-invasive ventilation. METHODS: The patients were divided into 2 groups: surfactant (n = 33) and control (n = 32). The subjects in the surfactant group received the inhaled surfactant at daily dose of 150-300 mg. The oxygenation parameters and several clinical outcomes were analyzed. RESULTS: On the 5 day of therapy, PaO2/FiO2 improved significantly in the surfactant group compared to the control group (184 (155-212) mmHg vs 150 (91-173) mmHg, p = 0.02). The inhaled surfactant significantly reduced the need for transfer of patients to intensive care units (24.2% vs 46.9%, p = 0.05) and invasive mechanical ventilation (18.2% vs 40.6%, p = 0.04). Even more, the nebulized surfactant shortened the length of non-invasive ventilation (7 (3-13) days vs 11 (5-22) days, p = 0.02) and time spent in hospital (18 (16-27) days vs 26 (21-31) days, p = 0.003) in patients suffering from COVID-19-linked ARDS. CONCLUSIONS: Our preliminary data provided indications that inhaled surfactant therapy may represent a promising option for patients with COVID-19-associated ARDS. However, larger clinical trials are crucially needed.

Pulmonary Hypertension in Acute and Chronic High Altitude Maladaptation Disorders.

Alveolar hypoxia is the most prominent feature of high altitude environment with well-known consequences for the cardio-pulmonary system, including development of pulmonary hypertension. Pulmonary hypertension due to an exaggerated hypoxic pulmonary vasoconstriction contributes to high altitude pulmonary edema (HAPE), a life-threatening disorder, occurring at high altitudes in non-acclimatized healthy individuals. Despite a strong physiologic rationale for using vasodilators for prevention and treatment of HAPE, no systematic studies of their efficacy have been conducted to date. Calcium-channel blockers are currently recommended for drug prophylaxis in high-risk individuals with a clear history of recurrent HAPE based on the extensive clinical experience with nifedipine in HAPE prevention in susceptible individuals. Chronic exposure to hypoxia induces pulmonary vascular remodeling and development of pulmonary hypertension, which places an increased pressure load on the right ventricle leading to right heart failure. Further, pulmonary hypertension along with excessive erythrocytosis may complicate chronic mountain sickness, another high altitude maladaptation disorder. Importantly, other causes than hypoxia may potentially underlie and/or contribute to pulmonary hypertension at high altitude, such as chronic heart and lung diseases, thrombotic or embolic diseases. Extensive clinical experience with drugs in patients with pulmonary arterial hypertension suggests their potential for treatment of high altitude pulmonary hypertension. Small studies have demonstrated their efficacy in reducing pulmonary artery pressure in high altitude residents. However, no drugs have been approved to date for the therapy of chronic high altitude pulmonary hypertension. This work provides a literature review on the role of pulmonary hypertension in the pathogenesis of acute and chronic high altitude maladaptation disorders and summarizes current knowledge regarding potential treatment options.