Development Of Pulmonary Arterial Hypertension Research Articles

Deficiency of the Deubiquitinase UCHL1 Attenuates Pulmonary Arterial Hypertension.

The ubiquitin-proteasome system regulates protein degradation and the development of pulmonary arterial hypertension (PAH), but knowledge about the role of deubiquitinating enzymes in this process is limited. UCHL1 (ubiquitin carboxyl-terminal hydrolase 1), a deubiquitinase, has been shown to reduce AKT1 (AKT serine/threonine kinase 1) degradation, resulting in higher levels. Given that AKT1 is pathological in pulmonary hypertension, we hypothesized that UCHL1 deficiency attenuates PAH development by means of reductions in AKT1. Tissues from animal pulmonary hypertension models as well as human pulmonary artery endothelial cells from patients with PAH exhibited increased vascular UCHL1 staining and protein expression. Exposure to LDN57444, a UCHL1-specific inhibitor, reduced human pulmonary artery endothelial cell and smooth muscle cell proliferation. Across 3 preclinical PAH models, LDN57444-exposed animals, Uchl1 knockout rats (Uchl1-/-), and conditional Uchl1 knockout mice (Tie2Cre-Uchl1fl/fl) demonstrated reduced right ventricular hypertrophy, right ventricular systolic pressures, and obliterative vascular remodeling. Lungs and pulmonary artery endothelial cells isolated from Uchl1-/- animals exhibited reduced total and activated Akt with increased ubiquitinated Akt levels. UCHL1-silenced human pulmonary artery endothelial cells displayed reduced lysine(K)63-linked and increased K48-linked AKT1 levels. Supporting experimental data, we found that rs9321, a variant in a GC-enriched region of the UCHL1 gene, is associated with reduced methylation (n=5133), increased UCHL1 gene expression in lungs (n=815), and reduced cardiac index in patients (n=796). In addition, Gadd45α (an established demethylating gene) knockout mice (Gadd45α-/-) exhibited reduced lung vascular UCHL1 and AKT1 expression along with attenuated hypoxic pulmonary hypertension. Our findings suggest that UCHL1 deficiency results in PAH attenuation by means of reduced AKT1, highlighting a novel therapeutic pathway in PAH.

Combined physical training protects the left ventricle from structural and functional damages in experimental pulmonary arterial hypertension

BackgroundUnder the adverse remodeling of the right ventricle and interventricular septum in pulmonary arterial hypertension (PAH) the left ventricle (LV) dynamics is impaired. Despite the benefits of combined aerobic and resistance physical trainings to individuals with PAH, its impact on the LV is not fully understood.ObjectiveTo test whether moderate-intensity combined physical training performed during the development of PAH induced by MCT in rats is beneficial to the LV’s structure and function.MethodsMale Wistar rats were divided into two groups: Sedentary Hypertensive Survival (SHS, n = 7); and Exercise Hypertensive Survival (EHS, n = 7) to test survival. To investigate the effects of combined physical training, another group of rats were divided into three groups: Sedentary Control (SC, n = 7); Sedentary Hypertensive (SH, n = 7); and Exercise Hypertensive (EH, n = 7). PAH was induced through an intraperitoneal injection of MCT (60 mg/kg). Echocardiographic evaluations were conducted on the 22nd day after MCT administration. Animals in the EHS and EH groups participated in a combined physical training program, alternating aerobic (treadmill running: 50 min, 60% maximum running speed) and resistance (ladder climbing: 15 climbs with 1 min interval, 60% maximum carrying load) exercises, one session/day, 5 days/week for approximately 4 weeks.ResultsThe physical training increased survival and tolerance to aerobic (i.e., maximum running speed) and resistance (i.e., maximum carrying load) exertions and prevented reductions in ejection fraction and fractional shortening. In addition, the physical training mitigated oxidative stress (i.e., CAT, SOD and MDA) and inhibited adverse LV remodeling (i.e., Collagen, extracellular matrix, and cell dimensions). Moreover, the physical training preserved the amplitude and velocity of contraction and hindered the reductions in the amplitude and velocity of the intracellular Ca2+ transient in LV single myocytes.ConclusionModerate-intensity combined physical training performed during the development of MCT-induced PAH in rats protects their LV from damages to its structure and function and hence increases their tolerance to physical exertion and prolongs their survival.

Lung capillary endothelium to arterial endothelium transition in pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) is characterized by a progressive increase of pulmonary vascular resistance and obliterative pulmonary vascular remodeling that result in right heart hypertrophy, failure, and premature death. The underlying mechanisms of loss of distal capillary endothelial cells (ECs) and obliterative vascular lesion formation remain unclear. Our recent single-cell RNA sequencing, spatial transcriptomics analysis, RNASCOPE, and immunostaining analysis showed that arterial ECs accumulation and loss of capillary ECs were evident in human PAH patients and pulmonary hypertension (PH) rodents. Pseudotime trajectory analysis of the single-cell RNA sequencing data suggest that lung capillary ECs transit to arterial ECs during the development of PH. Our study also identified CXCL12 as the marker for arterial ECs in PH. General capillary EC lineage tracing approach using Plvap-DreERT2;Tdtomato mice demonstrated that general capillary ECs gave rise to arterial ECs during PH development. Genetic deletion of HIF-2a or pharmacological inhibition of Notch4 normalized the arterial programming in PH. In conclusion, our study demonstrates that general capillary endothelium transits to arterial endothelium through the HIF-2a-Notch4 pathway during the development of PAH. Thus, targeting arterial EC transition might be a novel approach for treating PAH patients. This work was supported in part by NIH grant R00HL138278, R01HL158596, R01HL62794, R01HL169509, R01HL170096, AHA Career Development Award 20CDA35310084, The Cardiovascular Research and Education Foundation, Arizona Biomedical Research Centre funding (RFGA2022-01-06), and University of Arizona institution funding to Z.D. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Pathobiology of Pulmonary Arterial Hypertension

Abstract: Pulmonary Arterial Hypertension (PAH) is a progressive disease associated with occlusive pulmonary arterial remodeling of vessels < 500 μm for which there is no cure. Even in the era of PAH-specific combination therapies, aberrant lung pathology and progressive right ventricular (RV) dysfunction occur, culminating in a median survival of 6.2 years, according to the latest data in the treatment era. While better than a median survival from symptom onset of 2.8 years prior to PAH-specific therapies, it is still poor. Thus, there is an urgent need to move the opportunities forward for meaningful treatment strategies. Clearly, a better understanding of the highly complex pathobiology of PAH is needed if we are to achieve new and novel treatment strategies. This is especially so if we are to pursue a more personalized treatment approach to PAH in light of the multitude of pathobiological abnormalities described in PAH, which likely culminate in a final common pathway for PAH development. In this State-of-the-Art review, we provide comprehensive insights into the complex pathobiology of PAH to provide understanding and insights for the practicing clinician. We review the pathology of PAH and the cells involved and their impact in driving pathological abnormalities (pulmonary artery endothelial cells, smooth muscle cells, fibroblasts and pericytes) as well as the role of the extracellular matrix. Inflammation and immune dysfunction are considered important drivers of PAH and are comprehensively discussed. Another pathway relates to TGFβ/ bone morphogenic protein (BMP) imbalance, which is highlighted, as well as a new novel agent, sotatercept that impacts this imbalance. Genetic factors underlying heritable PAH (HPAH) are addressed, as well as epigenetic influences. Other important pathways highlighted include growth factor signaling, ion channels/channelopathy, hypoxia signaling pathways, and altered metabolism and mitochondrial dysfunction. We also address the “estrogen paradox”, whereby PAH is more common in women but more severe in men. The basis for drug-induced PAH is discussed, including the new methamphetamine epidemic. We briefly provide insights into DNA damage and senescence factors in pathobiology and highlight commonalities between PAH and cancer pathobiology. Furthermore, we provide concluding insights for the treating physician. In conclusion, we need to pose the right questions to motivate novel and effective treatment strategies for the management of PAH based on pathobiological principles and understanding.

Neutrophil extracellular traps promote proliferation of pulmonary smooth muscle cells mediated by CCDC25 in pulmonary arterial hypertension

BackgroundPrevious studies have indicated that neutrophil extracellular traps (NETs) play a pivotal role in pathogenesis of pulmonary arterial hypertension (PAH). However, the specific mechanism underlying the impact of NETs on pulmonary artery smooth muscle cells (PASMCs) has not been determined. The objective of this study was to elucidate underlying mechanisms through which NETs contribute to progression of PAH.MethodsBioinformatics analysis was employed in this study to screen for potential molecules and mechanisms associated with occurrence and development of PAH. These findings were subsequently validated in human samples, coiled-coil domain containing 25 (CCDC25) knockdown PASMCs, as well as monocrotaline-induced PAH rat model.ResultsNETs promoted proliferation of PASMCs, thereby facilitating pathogenesis of PAH. This phenomenon was mediated by the activation of transmembrane receptor CCDC25 on PASMCs, which subsequently activated ILK/β-parvin/RAC1 pathway. Consequently, cytoskeletal remodeling and phenotypic transformation occur in PASMCs. Furthermore, the level of NETs could serve as an indicator of PAH severity and as potential therapeutic target for alleviating PAH.ConclusionThis study elucidated the involvement of NETs in pathogenesis of PAH through their influence on the function of PASMCs, thereby highlighting their potential as promising targets for the evaluation and treatment of PAH.

Activation of CaMKII/HDAC4 by SDF1 contributes to pulmonary arterial hypertension via stabilization Runx2

Stromal derived factor 1 (SDF1) has been shown to be involved in the pathogenesis of pulmonary artery hypertension (PAH). However, the detailed molecular mechanisms remain unclear. To address this, we utilized primary cultured rat pulmonary artery smooth muscle cells (PASMCs) and monocrotaline (MCT)-induced PAH rat models to investigate the mechanisms of SDF1 driving PASMCs proliferation and pulmonary arterial remodeling. SDF1 increased runt-related transcription factor 2 (Runx2) acetylation by Calmodulin (CaM)-dependent protein kinase II (CaMKII)-dependent HDAC4 cytoplasmic translocation, elevation of Runx2 acetylation conferred its resistance to proteasome-mediated degradation. The accumulation of Runx2 further upregulated osteopontin (OPN) expression, finally leading to PASMCs proliferation. Blocking SDF1, suppression of CaMKII, inhibition the nuclear export of HDAC4 or silencing Runx2 attenuated pulmonary arterial remodeling and prevented PAH development in MCT-induced PAH rat models. Our study provides novel sights for SDF1 induction of PASMCs proliferation and suggests that targeting SDF1/CaMKII/HDAC4/Runx2 axis has potential value in the management of PAH.

Mechanistic Investigation of Calcium Channel Regulation-Associated Genes in Pulmonary Arterial Hypertension and Signatures for Diagnosis.

Pulmonary arterial hypertension (PAH) is a severe cardiopulmonary disorder with complex causes. Calcium channel blockers have long been used in its treatment. Our study aimed to validate experimental results showing increased calcium ion concentration in PAH patients. We investigated the impact of genes related to calcium channel regulation on PAH development and developed an accurate diagnostic model. Clinical trial data from serum of 18 healthy individuals and 18 patients with PAH were retrospectively analyzed. Concentrations of calcium and potassium ions were determined and compared. Datasets were retrieved, selecting genes associated with calcium ion release. R packages processed the datasets, filtering 174 common genes, and conducting Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses. Six hub genes were identified, and nomogram and logistic regression prediction models were constructed. Random forest filtered cross genes, and a diagnostic model was developed and validated using an artificial neural network. The 174 intersection genes related to calcium ions showed significant correlations with biological processes, cellular components, and molecular functions. Six key genes were obtained by constructing a protein-protein interaction network. A diagnostic model with high accuracy (> 90%) and diagnostic capability (AUC = 0.98) was established using a neural network algorithm. This study validated the experimental results, identified key genes associated with calcium ions, and developed a highly accurate diagnostic model using a neural network algorithm. These findings provide insights into the role of calcium release genes in PAH and demonstrate the potential of the diagnostic model for clinical application. However, due to limitations in sample size and a lack of prognosis data, the regulatory mechanisms of calcium ions in PAH patients and their impact on the clinical prognosis of PAH patients still need further exploration in the future.

Causal effect of interleukin (IL)-6 on blood pressure and hypertension: A mendelian randomization study.

To examine whether circulating interleukin-6 (IL-6) levels (CirIL6) have a causal effect on blood pressure using Mendelian randomization (MR) methods. We used data from genome-wide association studies (GWAS) of European ancestry to obtain genetic instruments for circulating IL-6 levels and blood pressure measurements. We applied several robust MR methods to estimate the causal effects and to test for heterogeneity and pleiotropy. We found that circulating IL-6 had a significant positive causal effect on systolic blood pressure (SBP) and pulmonary arterial hypertension (PAH), but not on diastolic blood pressure (DBP) or hypertension. We found that as CirIL6 genetically increased, SBP increased using Inverse Variance Weighted (IVW) method (for ukb-b-20175, β = 0.082 with SE = 0.032, P = 0.011; for ukb-a-360, β = 0.075 with SE = 0.031, P = 0.014) and weighted median (WM) method (for ukb-b-20175, β = 0.061 with SE = 0.022, P = 0.006; for ukb-a-360, β = 0.065 with SE = 0.027, P = 0.014). Moreover, CirIL6 may be associated with an increased risk of PAH using WM method (odds ratio (OR) = 15.503, 95% CI, 1.025-234.525,P = 0.048), but not with IVW method. Our study provides novel evidence that circulating IL-6 has a causal role in the development of SBP and PAH, but not DBP or hypertension. These findings suggest that IL-6 may be a potential therapeutic target for preventing or treating cardiovascular diseases and metabolic disorders. However, more studies are needed to confirm the causal effects of IL-6 on blood pressure and to elucidate the underlying mechanisms and pathways.

Activation of the p62-Keap1-Nrf2 pathway improves pulmonary arterial hypertension in MCT-induced rats by inhibiting autophagy.

Autophagy is implicated in the pathogenesis of pulmonary arterial hypertension (PAH). We aimed to investigate whether the p62-Keap1-Nrf2 pathway affects the development of PAH by mediating autophagy. A PAH rat model was established using monocrotaline (MCT). Pulmonary artery smooth muscle cells (PASMCs) were extracted, and the changes in proliferation, migration, autophagy, and oxidative stress were analyzed following overexpression or knockdown of p62. The impact of p62 on the symptoms of PAH rats was assessed by the injection of an adenovirus overexpressing p62. We found that the knockdown of p62 increased the proliferation and migration of PASMCs, elevating the oxidative stress of PASMCs and upregulating gene expression of NADPH oxidases. Co-IP assay results demonstrated that p62 interacted with Keap1. p62 knockdown enhanced Keap1 protein stability and Nrf2 ubiquitination. LC3II/I and ATG5 were expressed more often when p62 was knocked down. Treating with an inhibitor of autophagy reversed the impact of p62 knockdown on PASMCs. Nrf2 inhibitor treatment reduced the expression of Nrf2 and p62, while increasing the expression of Keap1, LC3II/I, and ATG5 in PASMCs. However, overexpressing p62 diminished mRVP, SPAP, and Fulton index in PAH rats and attenuated pulmonary vascular wall thickening. Overexpression of p62 also decreased the expression of Keap1, LC3II/I, and ATG5 and increased the nuclear expression of Nrf2 in PAH rats. Importantly, overexpression of p62 reduced oxidative stress and the NADPH oxidase expression in PAH rats. Overall, activation of the p62-Keap1-Nrf2 positive feedback signaling axis reduces the proliferation and migration of PASMCs and alleviates PAH by inhibiting autophagy and oxidative stress.

Nailfold capillaroscopy abnormalities and pulmonary hypertension in mixed connective tissue disease and systemic sclerosis patients.

Pulmonary arterial hypertension (PAH) represents an important vascular complication of mixed connective tissue disease (MCTD) and systemic sclerosis (SSc). Microvascular involvement in these diseases can be investigated by means of nailfold capillaroscopy (NFC). Microvascular involvement detected in the nailfold bed is the mirror of the microvascular damage occurring in the entire body, further indicating the involvement of the target organs. The aim of this study was to evaluate the microvascular involvement in MCTD patients with or without PAH, compared to that found in SSc patients with or without PAH. This cross-sectional study was performed in the Department of Internal Medicine and Department of Rheumatology, Timișoara, Romania, during the time period between January 2017 and December 2022, on a group of 26 patients with MCTD and 26 SSc patients. Antinuclear antibodies, anti-U1-RNP, anti-Scl 70, anti-centromere, anti-cardiolipin antibodies (aCL) (IgM, IgG), anti-β2-glycoprotein I (aβ2GPI) (IgM, IgG) antibodies, and lupus anticoagulant (LAC) were determined in both the groups. PAH was evaluated through cardiac ultrasonography, determining the sPAP (systolic pulmonary artery pressure). Nailfold capillaroscopy was performed using a USB Digital Microscope and 2.0-megapixel digital camera recording capillaries density, giant capillaries, enlarged capillaries, capillaries hemorrhages, avascular areas, ramified/bushy capillaries scores. Data were recorded and presented as mean ± standard deviation. Statistical analyses were performed using the Student's t-test, ANOVA test, and Pearson's correlation. Differences were considered statistically significant if p-value < 0.05. Among the MCTD patients, PAH was identified in 12 patients (46.15%), while among the SSc patients PAH was identified in 14 patients (53.84%). Development of PAH in MCTD patients was associated with lower capillaries density (p-value < 0.00001), higher scores of giant capillaries, ramified/bushy capillaries, and capillary hemorrhages (p-value < 0.00001, for each of them). Anti-U1-RNP, aCL, aβ2GPI antibodies and LAC were also found to be involved in PAH-associated MCTD development. Unlike MCTD patients, SSc patients with PAH presented with lower capillaries density and ramified/bushy capillaries scores (p-value < 0.05). The MCTD patients who presented significant NFC abnormalities (especially active and late scleroderma-like capillaroscopic pattern) are prone to PAH development. Capillary density reduction is the most important factor associated with the occurrence of PAH. Differences in NFC findings (especially capillary density and ramified/bushy capillaries) were detected among patients with MCTD and SSc having PAH.

RAB7 deficiency impairs pulmonary artery endothelial function and promotes pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is a devastating and progressive disease with limited treatment options. Endothelial dysfunction plays a central role in the development and progression of PAH, yet the underlying mechanisms are incompletely understood. The endosome-lysosome system is important to maintain cellular health, and the small GTPase RAB7 regulates many functions of this system. Here, we explored the role of RAB7 in endothelial cell (EC) function and lung vascular homeostasis. We found reduced expression of RAB7 in ECs from patients with PAH. Endothelial haploinsufficiency of RAB7 caused spontaneous pulmonary hypertension (PH) in mice. Silencing of RAB7 in ECs induced broad changes in gene expression revealed via RNA-Seq, and RAB7-silenced ECs showed impaired angiogenesis and expansion of a senescent cell fraction, combined with impaired endolysosomal trafficking and degradation, suggesting inhibition of autophagy at the predegradation level. Furthermore, mitochondrial membrane potential and oxidative phosphorylation were decreased, and glycolysis was enhanced. Treatment with the RAB7 activator ML-098 reduced established PH in rats with chronic hypoxia/SU5416. In conclusion, we demonstrate for the first time to our knowledge the fundamental impairment of EC function by loss of RAB7, causing PH, and show RAB7 activation to be a potential therapeutic strategy in a preclinical model of PH.

The prognostic value of prognostic nutritional index in postoperative onset of PAH in children with isolated VSD: a prospective cohort study based on propensity score matching analysis.

The mechanism of pulmonary arterial hypertension (PAH) after surgery/intervention for isolated venticlular septal defect (VSD) in children is unknown. Reliable prognostic indicators for predicting postoperative PAH are urgently needed. Prognostic nutration index (PNI) is widely used to predict postoperative complications and survival in adults, but it is unclear whether it can be used as an indicator of prognosis in children. A total of 251 children underwent VSD repair surgery or interventional closure in Hunan Children's Hospital from 2020 to 2023 were collected. A 1:1 propensity score matching (PSM) analysis was performed using the nearest neighbor method with a caliper size of 0.2 Logistics regression analysis is used to examine factors associated with the development of PAH. The cut-off value for PNI was determined as 58.0. After 1:1 PSM analysis, 49 patients in the low PNI group were matched with high PNI group. Children in the low PNI group had higher risk of postoperative PAH (P = 0.002) than those in the high PNI group. Multivariate logistics regression analysis showed that PNI (RR: 0.903, 95% CI: 0.816-0.999, P = 0.049) and tricuspid regurgitation velocity (RR: 4.743, 95% CI: 1.131-19.897, P = 0.033) were independent prognostic factors for the development of PAH. PNI can be used as a prognostic indicator for PAH development after surgery/intervention in children with isolated VSD.

Анализ роли нейтрофилов в развитии воспаления в монокроталининдуцированной легочной артериальной гипертензии

Pulmonary arterial hypertension (PAH) is a clinical syndrome with dismal outcome. This disease is associated with endothelial dysfunction or damage, pulmonary vascular remodeling, and increased vascular resistance and pulmonary artery blood pressure. All this provokes right ventricular failure and pulmonary edema, which leads to premature death of patients. PAH is accompanied by the inflammatory process deve­lopment, one of the main causes of which is the increased production of reactive oxygen and chlorine species by stimulated neutrophils. The purpose of our study was to study the role of these cells in the oxidative stress formation in experimental model of PAH. The studies were conducted on outbred white rats. Modeling of PAH was carried out by subcutaneous administration of a monocrotaline solution at a dose of 60 mg/kg. The control group consisted of 10 healthy animals. After monocrotaline injection, PAH animals were divided into 4 groups: 10 rats in the 2-week group, 10 rats in the 4-week group, 14 rats in the 6-week group, and 20 rats in the 8-week group. After collecting peripheral blood and isolating neutrophils, reactive oxygen and chlorine species production, secretory degranulation and the unbound calcium ions concentration in the cytosol were determined using spectral analysis methods. The results were compared with corresponding data obtained for 10 healthy animals. It has been established that the development of PAH in rats over 8 weeks is accompanied by neutrophil priming and oxidative stress formation as a result of “respiratory burst” mechanisms modification. Changes in reactive oxygen and chlorine species production, disruption of Ca2+-dependent intracellular signaling involving cyclooxygenase 1/2 and phosphatidylinositol 3-kinase, and increased secretion of myeloperoxidase from neutrophils were revealed.

Evaluation of Risk Factors for Development of Pulmonary Hypertension in Patients with End Stage Renal Disease Undergoing Hemodialysis

Background: Pulmonary arterial hypertension (PAH) has recently been identified as a common complication in patients with end-stage renal disease (ESRD) who are undergoing hemodialysis (HD) or peritoneal dialysis (PD). The risk factors for the development of PAH in those patients are not well understood. Objective: To investigate the risk factors for the development of PAH in patients with ESRD on HD. Patients and Methods: This is a hospital-based cross-sectional study of 50 ESRD patients undergoing HD. The pulmonary artery systolic pressure (PASP) was measured using echocardiography. PASP ˃ 25 mmHg at rest was defined as PAH. As a result, patients were divided into two groups: those who had PAH and those who did not. Each patient's demographic, biochemical, and echographic findings were documented. Results: Out of 50 patients, 19 (38%) had PAH, while the remaining 31 (62%) had normal PASP. In multivariate analysis, HD duration &gt; 3.4 years (OR= 2.13, 95%CI=1.45-31.38, p= 0.025), hypertension as a cause of ESRD (OR=6.12, 95%CI=1.4-26.77, p=0.031), hemoglobin (Hb) ≤ 10.0 g/dl (OR= 4.35, 95%CI=1.88-9.84, p= 0.018), and left ventricular ejection fraction (LVEF)≤ 55% (OR= 6.75, 95%CI=1.87-23.74, p=0.021) were independent factors associated with PAH. PASP had a significant positive correlation with the rate of fistula flow (r= 0.295, p= 0.038) and E/A ratio (r= 0.368, p= 0.008), but a significant negative correlation with LVEF (r= -0.345, p= 0.014). PASP had a positive significant correlation with each of rate of fistula flow (r= 0.295, p= 0.038) and E/A ratio (r= 0.368, p= 0.008), while it has a negative significant correlation with LVEF (r= -0.345, p= 0.014). Conclusion: Longer duration of HD, hypertensive nephropathy as a cause of ESRD, Hb≤ 10 g/dl, and LVEF ≤55% are among the demographic, biochemical, and clinical factors associated with the development of PAH in patients with ESRD under HD. The PASP has a positive correlation with fistula flow rate and E/A ratio and a negative correlation with LVEF. Keywords: Pulmonary arterial hypertension, End stage renal disease, Hemodialysis.

Gut-Lung Axis in Focus: Deciphering the Impact of Gut Microbiota on Pulmonary Arterial Hypertension.

Recent advancements in the understanding of pulmonary arterial hypertension (PAH) have highlighted the significant role of the gut microbiota (GM) in its pathogenesis. This comprehensive review delves into the intricate relationship between the GM and PAH, emphasizing the influence of gut microbial composition and the critical metabolites produced. We particularly focus on the dynamic interaction between the gut and lung, examining how microbial dysbiosis contributes to PAH development through inflammation, altered immune responses, and changes in the gut-lung axis. Noteworthy findings include variations in the ratios of key bacterial groups such as Firmicutes and Bacteroidetes in PAH and the pivotal roles of metabolites like trimethylamine N-oxide (TMAO), short-chain fatty acids (SCFAs), and serotonin in the disease's progression. Additionally, the review elucidates potential diagnostic biomarkers and novel therapeutic approaches, including the use of probiotics and fecal microbiota transplantation, which leverage the gut microbiota for managing PAH. This review encapsulates the current state of research in this field, offering insights into the potential of gut microbiota modulation as a promising strategy in PAH diagnosing and treatment.

Modulating NO-GC Pathway in Pulmonary Arterial Hypertension.

The pathogenesis of complex diseases such as pulmonary arterial hypertension (PAH) is entirely rooted in changes in the expression of some vasoactive factors. These play a significant role in the onset and progression of the disease. Indeed, PAH has been associated with pathophysiologic alterations in vascular function. These are often dictated by increased oxidative stress and impaired modulation of the nitric oxide (NO) pathway. NO reduces the uncontrolled proliferation of vascular smooth muscle cells that leads to occlusion of vessels and an increase in pulmonary vascular resistances, which is the mainstay of PAH development. To date, two classes of NO-pathway modulating drugs are approved for the treatment of PAH: the phosphodiesterase-5 inhibitors (PD5i), sildenafil and tadalafil, and the soluble guanylate cyclase activator (sGC), riociguat. Both drugs provide considerable improvement in exercise capacity and pulmonary hemodynamics. PD5i are the recommended drugs for first-line PAH treatment, whereas sGCs are also the only drug approved for the treatment of resistant or inoperable chronic thromboembolic pulmonary hypertension. In this review, we will focus on the current information regarding the nitric oxide pathway and its modulation in PAH.

Endothelial cell clonality, heterogeneity and dysfunction in pulmonary arterial hypertension.

Our understanding of the pathophysiology of pulmonary arterial hypertension (PAH) has evolved over recent years, with the recognition that endothelial cell (EC) dysfunction and inflammation play an integral role in the development of this disease. ECs within the pulmonary vasculature play a unique role in maintaining vascular integrity and barrier function, regulating gas exchange, and contributing to vascular tone. Using single-cell transcriptomics, research has shown that there are multiple, unique EC subpopulations with different phenotypes. In response to injury or certain stressors such as hypoxia, there can be a dysregulated response with aberrant endothelial injury repair involving other pulmonary vascular cells and even immune cells. This aberrant signaling cascade is potentially a primary driver of pulmonary arterial remodeling in PAH. Recent studies have examined the role of EC clonal expansion, immune dysregulation, and genetic mutations in the pathogenesis of PAH. This review summarizes the existing literature on EC subpopulations and the intricate mechanisms through which ECs develop aberrant physiologic phenotypes and contribute to PAH. Our goal is to provide a framework for understanding the unique pulmonary EC biology and pathophysiology that is involved in the development of PAH.

Abstract 17430: Targeting DNA Hypermethylation With Hypomethylating Drugs: Implications for Pulmonary Arterial Hypertension

Introduction: Pulmonary arterial hypertension (PAH) is a lethal vasculopathy characterized by sustained vasoconstriction and remodeling of pulmonary arteries. Excessive proliferation and reduced apoptosis of pulmonary artery smooth muscle cells (PASMCs) contribute to pulmonary artery remodeling, with limited efficacy of current pharmacotherapies. Increased DNA methylation (DNAm) has been implicated in pro-proliferative phenotypes and disease progression in cancer and cardiovascular diseases. Hypothesis: We hypothesized that increased DNAm contributes to the pro-survival phenotype of PAH-PASMCs and that targeting DNA methylation could improve PAH development. Methods and Results: Our study revealed a global increase of DNAm in remodeled pulmonary arteries of human PAH patients and three preclinical models (namely monocrotaline, sugen hypoxia, Fawn hooded rats, p&lt;0.05, immunofluorescence [IF] 5mc/SMA). In vitro, PAH-PASMCs exhibited elevated DNAm levels (IF 5mc, p&lt;0.05), increased expression of DNA methyltransferases (DNMT1, DNMT3a; Western blot [WB], p&lt;0.05), and decreased expression of DNA demethylase (TET2; WB, p&lt;0.05). Consequently, hypomethylating drug Zebularine demonstrated dose-dependent reductions in PAH-PASMC proliferation (WB for surviving, plk1, MCM2; IF ki67, p&lt;0.05) and increased apoptosis (WB for Bax/BCL2; IF annexinV, p&lt;0.05,). Transcriptomic and gene ontology analyses indicated that these effects likely involve the downregulation of biological processes related to the cell cycle and mitosis. In vivo treatment with Zebularine (10 mg/kg, 2 weeks, intraperitoneal injection) improved PAH hemodynamics (e.g., decreased RVSP, vascular resistances, p&lt;0.05), decreased PASMC methylation (IF 5mc/SMA, p&lt;0.05) and proliferation (IF ki67/SMA, p&lt;0.05), and mitigated adverse pulmonary vascular remodeling in two rodent models of the disease (monocrotaline and sugen hypoxia). Statistics: unpaired t test (2 groups), ANOVA (&gt; 2 groups). Conclusions: Our findings demonstrate an association between increased DNA methylation and PAH development. Targeting DNAm represents a potential therapeutic avenue for PAH, as evidenced by the beneficial effects of Zebularine in improving PAH-related phenotypes.

Abstract 15967: Unravelling the Role of Deoxyhypusine Synthase-Mediated Hypusination of eIF5A in Pulmonary Arterial Hypertension

Introduction: Pulmonary Arterial Hypertension (PAH) is characterized by progressive distal pulmonary arteries (PAs) obstruction leading to heart failure and death. PA smooth muscle cells (PASMCs) from PAH patients display a “cancer-like” phenotype that contributes to PA remodeling. Appreciation of the pivotal role of translational control in hyperproliferating diseases is steadily increasing. In this regard, eukaryotic translation initiation factor 5A (eIF5A) was shown to provide cancer cells with a competitive advantage by increasing translation of mRNAs with oncogenic proprieties. Strikingly, eIF5A is the only protein containing the unique spermidine-derived amino acid hypusine required for its function. Hypusine formation is catalyzed by the sequential actions of deoxyhypusine synthase (DHPS) and deoxyhypusine hydrolase (DOHH). We hypothesized that increased Hyp eIF5A in PAH-PASMCs is required to promote translational efficiency of a set of factors conferring a higher survival and proliferative capacity, leading to pulmonary vascular remodeling. Methods/Results: As assessed by LC-MSMS and WB, expression levels of DHPS, DOHH and both total and hypusinated forms of eIF5A were found increased in PAs and PASMCs from PAH patients and animal models. In vitro , both molecular and pharmacological inhibition of DHPS and DOHH significantly attenuated PAH-PASMCs survival (WB Survivin; Annexin V labeling) and proliferation (WB MCM2, PLK1; Ki67 labeling and EdU incorporation). Hypusine signaling promoted the expression of a broad array of proteins involved in oxidative phosphorylation, supporting the bioenergetic requirements of cell survival and proliferation (LC-MSMS, WB, Seahorse). In vivo , smooth muscle cells-targeted inactivation of one allele of Dhps conferred partial protection against the development of Sugen/Hypoxia (Su/Hx)-induced PAH in mice. Accordingly, pharmacological inhibition of DHPS using GC7 improved hemodynamics (RVSP, mPAP, CO) and vascular remodeling (EVG) in monocrotaline and Su/Hx rats with established PAH. Conclusion: We showed for the first time that hypusine signaling is implicated in PAH development and represents a new promising therapeutic target.

Abstract 14829: Inflammatory Cells and Cytokines Are Increased in Mice With Pulmonary Arterial Hypertension Associated With Hematopoietic Dnmt3a Depletion

Background: Pulmonary Arterial Hypertension (PAH) is characterized by pulmonary vascular remodeling of the precapillary pulmonary arteries. Increasing evidence suggests that inflammation plays an essential role in PAH development. Understanding the involvement of immunity in this context can provide valuable insights into the pathogenesis of PAH. DNA Methyltransferase 3A ( DNMT3A ) mutations can lead to altered DNA methylation patterns in immune cells, potentially affecting the expression of immune-regulatory genes. Here, we investigate the role of inflammation in PAH driven by hematopoietic Dnmt3a depletion in mice. Methods: We used male and female conditional, Vav-Cre-driven hematopoietic Dnmt3a knockout mice ( Dnmt3a –/– ) and compared them to controls (n=3/group). Lung tissue and plasma were collected. Single-cell suspensions of digested lungs were stained to detect total leukocytes (CD45+), T cells (CD3+), B cells (CD19+), macrophages (Ly6G-, F4/80+), and neutrophils (Ly6G+, F4/80-) via flow cytometry. Plasma samples were screened for 32 distinct cytokines using a cytokine array. Results: Dnmt3a –/– mice spontaneously developed PAH at 4.5 months. PAH was associated with a 19% increase in total CD45+ leukocytes in the lungs compared to controls. T cells, and B cells were increased in Dnmt3a –/– mice compared to controls (7.9% and 3.1%, respectively). In contrast, there was no difference in lung neutrophil count. Two subsets of macrophages were observed based on granularity. Compared to controls, Dnmt3a –/– mice had a 29% increase in agranular macrophages and a 19.7% increase in granular macrophages. IL-13, known for its role in alternative monocyte/macrophage activation in PAH patients associated with systemic sclerosis (SSc-PAH), was significantly increased in the plasma of Dnmt3a –/– mice compared to controls (1.8 times higher, p=0.02). IL-12p40, a chemoattractant for macrophages, showed a trend towards being increased in Dnmt3a –/– mice compared to controls (5.7 times higher, p=0.14). Conclusion: Deletion of DNMT3A in the hemopoietic system triggers an inflammatory form of PAH in mice, associated with increased macrophages, B- and T-cells in the lung. IL-13 may be a potential diagnostic marker in PAH.