Pulmonary Arterial Remodeling Research Articles

NCAM1 modulates the proliferation and migration of pulmonary arterial smooth muscle cells in pulmonary hypertension

BackgroundPulmonary hypertension (PH) is a malignant vascular disease characterized by pulmonary arterial remodeling. Neural cell adhesion molecule 1 (NCAM1) is a cell surface glycoprotein that is involved in a variety of diseases, including cardiovascular disease. However, the role of NCAM1 in PH remains underexplored.MethodsPulmonary hypertension models were established using monocrotaline in rats and hypoxia in mice. NCAM1 protein levels in plasma from patients and rats were measured by ELISA. Expression of NCAM1 in rat lung tissues were evaluated using qRT-PCR, Western blotting, and immunofluorescence. The effects of NCAM1 on rat pulmonary artery smooth muscle cells were studied by stimulating these cells with PDGF-BB.ResultsElevated levels of NCAM1 protein and mRNA were observed in both PH patients and monocrotaline-induced PH rats. NCAM1 knockdown ameliorated hypoxia-induced PH, highlighting its role in pulmonary artery remodeling. In PASMCs, NCAM1 expression was upregulated by PDGF-BB stimulation, enhancing cell proliferation and migration. This effect was attenuated by NCAM1 knockdown but partially restored by an ERK1/2 pathway activator (tert-butylhydroquinone, TBHQ), suggesting NCAM1’s involvement in PASMC dynamics through the ERK1/2 signaling pathway.ConclusionOur findings confirm the role of NCAM1 in pulmonary arterial hypertension and demonstrate its promotion of PASMC proliferation and migration through the ERK1/2 signaling pathway.

Endothelial FUNDC1 Deficiency Drives Pulmonary Hypertension.

Pulmonary hypertension (PH) is associated with endothelial dysfunction. However, the cause of endothelial dysfunction and its impact on PH remain incompletely understood. We aimed to investigate whether the hypoxia-inducible FUNDC1 (FUN14 domain-containing 1)-dependent mitophagy pathway underlies PH pathogenesis and progression. We first analyzed FUNDC1 protein levels in lung samples from patients with PH and animal models. Using rodent PH models induced by HySu (hypoxia+SU5416) or chronic hypoxia, we further investigated PH pathogenesis and development in response to global and cell-type-specific Fundc1 loss/gain-of-function. We also investigated the spontaneous PH in mice with inducible loss of endothelial Fundc1. In addition, histological, metabolic, and transcriptomic studies were performed to delineate molecular mechanisms. Finally, findings were validated in vivo by compound deficiency of HIF2α (hypoxia-inducible factor 2α; Epas1) and pharmacological intervention. FUNDC1 protein levels were reduced in PH lung vessels from clinical subjects and animal models. Global Fundc1 deficiency exacerbated PH, while its overexpression is protective. The effect of FUNDC1 was mediated by endothelial cells rather than smooth muscle cells. Further, inducible loss of endothelial Fundc1 in postnatal mice was sufficient to cause PH spontaneously, whereas augmenting endothelial Fundc1 protected against PH before and after the onset of disease. Mechanistically, Fundc1 deficiency impaired basal mitophagy in endothelial cells, leading to the accumulation of dysfunctional mitochondria, metabolic reprogramming toward aerobic glycolysis, pseudohypoxia, and senescence, likely via a mtROS-HIF2α signaling pathway. Subsequently, Fundc1-deficient endothelial cells increased IGFBP2 (insulin-like growth factor-binding protein 2) secretion that drove pulmonary arterial remodeling to instigate PH. Finally, proof-of-principle in vivo studies showed significant efficacy on PH amelioration by targeting endothelial mitophagy, pseudohypoxia, senescence, or IGFBP2. Collectively, we show that FUNDC1-mediated basal mitophagy is critical for endothelial homeostasis, and its disruption instigates PH pathogenesis. Given that similar changes in FUNDC1 and IGFBP2 were observed in PH patients, our findings are of significant clinical relevance and provide novel therapeutic strategies for PH.

Abstract 4119280: The role of colony-stimulating factor 1 receptor and perivascular macrophages in pulmonary arterial hypertension

Background: Pulmonary arterial hypertension (PAH) is a life-threatening disease caused by pulmonary artery remodeling, inflammation, and altered immune responses. The proliferation of pulmonary arterial smooth muscle cells and the accumulation of perivascular macrophages are crucial factors for the development of pulmonary artery remodeling. However, the mechanism for the progression of pulmonary artery remodeling through the perivascular macrophages has not been fully investigated. In this study, we examined the significance of colony-stimulating factor 1 receptor (CSF1R), which regulates the differentiation and proliferation of macrophages in PAH. Methods and Results: The numbers of pulmonary arterial perivascular CSF1R-positive cells significantly increased in lungs from PAH patients, compared to controls. The CSF1R phosphorylation on whole lungs was significantly increased in monocrotaline (MCT)-induced PAH rats and SU5416/Hypoxia (SuHx)-induced PAH mice. Next, we evaluated the therapeutic potential of a CSF1R inhibitor or anti-CSF1R antibody in vivo . Blocking CSF1R with a CSF1R inhibitor or anti-CSF1R antibody significantly decreased right ventricular systolic pressure, right ventricular fibrosis, perivascular CD68-positive macrophages and Arg1-positive M2 macrophages in PAH rats and mice. CSF1R knockdown using antisense oligonucleotide in the lung significantly decreased right ventricular systolic pressure in SuHx-induced PAH mice. Next, we focused on C-C motif chemokine ligand 2 (CCL2) whose gene expression is elevated in macrophages of PAH model. The culture with a conditioned medium from lung-derived M2 macrophages accelerated the proliferation of pulmonary arterial smooth muscle cells, while adding a CCL2 neutralizing antibody into the conditioned medium inhibited their proliferation. Conclusion: CSF1R has a critical role in the proliferation of pulmonary arterial perivascular macrophages and the progression of PAH, and M2-derived CCL2 contributed to the proliferation of pulmonary arterial smooth muscle cells. CSF1R is a novel therapeutic target in PAH.

Abstract 4138731: Clonal Hematopoiesis of Indeterminant Potential is Associated with Pulmonary Arterial Hypertension

Background: Pulmonary Arterial Hypertension (PAH) is a fatal cardiopulmonary disorder characterized by adverse vascular remodelling of small pulmonary arteries, which increases resistance and demand on the right ventricle (RV), ultimately leading to RV fibrosis and failure. Emerging evidence suggests that inflammation plays a crucial role in PAH pathogenesis. Here, we study Clonal Hematopoiesis of Indeterminate Potential (CHIP), a disorder characterized by somatic mutations in genes such as DNMT3A and TET2 , that promotes an inflammatory phenotype, in the development of PAH. Methods: We performed deep, targeted panel sequencing on 1659 PAH patients from the PAH Biobank and 3644 controls (3401 Vanderbilt University Biorepository patients and 243 participants recruited by the Queen’s Genomics Centre at Ongwanada). Variants in known CHIP genes expressed at a variant allele frequency (VAF) > 2% were classified as CHIP. Genes with a VAF > 25% and <50% were considered CHIP if located in reported CHIP hotspot regions. The prevalence of CHIP mutations was compared between controls and PAH patients. Logistic regression was used for statistical analysis. Results: We identified 242 CHIP variants in the 1659 PAH patients. DNMT3A was the most mutated gene (116/242 = 48%), followed by TET2 (46/242=19%). After adjusting for age and sex, we identified a significant association between all CHIP variants combined and PAH (OR: 23.35 [7.67, 71.03]; p<0.0001). Further, DNMT3A CHIP (OR: 25.44 [5.37, 120.40]; p<0.0001), non- DNMT3A CHIP (OR: 26.80 [5.56, 129.23]; p<0.0001) and TET2 CHIP (OR: 13.69 [1.29, 145.30]; p=0.03) were all associated with PAH. A higher proportion of PAH patients under the age of 70 were found to have CHIP variants compared to controls and this was significant for individuals between 60-69 years of age (p=0.037). While there was a 3.8:1 female-to-male ratio of PAH patients, there was a 5.8:1 female-to-male preponderance of DNMT3A variants (p=0.039) (4.3:1 for all CHIP; p=ns), suggesting DNMT3A variants are more common in female patients. The presence of CHIP variants had no significant effect on PAH patient 10-year survival. Conclusion: This research reveals a significant association between CHIP variants, specifically in DNMT3A and TET2, and PAH. The prevalence of CHIP is higher in PAH patients aged 40 to 70 compared to controls and is more common in female patients. These findings suggest that CHIP is a significant risk factor for the development of PAH.

Pulmonary hypertension in patients carrying FLNA loss-of-function variants.

Pulmonary hypertension (PH) is an unusual complication of X-linked disease caused by loss-of-function (LOF) variants in the filamin A (FLNA) gene. Patients with FLNA LOF may also present dysmorphic facial features, aortic dilation, thrombopenia, and periventricular nodular heterotopia (PVNH). We reported clinical, functional, radiologic, and hemodynamic characteristics of patients with FLNA LOF variants and PH from the French PH Network. Nine patients were identified with a female to male ratio of 8:1. PH was diagnosed at a median age of 36 [0-69] years. Associated conditions included epilepsy (n=5), PVNH (n=7), valvular heart disease (n=8), congenital heart diseases (n=4), thrombocytopenia (n=4), and hyperlaxity (n=4). Right heart catheterisation confirmed moderate-to-severe precapillary PH with a median mPAP of 33 [22-49] mmHg and PVR of 4.7 [2.4-8.0] WU. The DLCO was markedly decreased (48 [22-64] %pred) and five patients had obstructive ventilatory disorder. High-resolution CT showed heterogeneous parenchyma (n=8), emphysema (n=3), presence of a peripheral hyperclear band (n=3) and aortic ectasia (n=4). Pathologic assessment available in one patient revealed significant remodelling of small pulmonary arteries, interstitial edema, and irregular alveoli shapes. During follow-up, three patients died, including two from right heart failure. No patient died from aortic rupture. Precapillary PH, likely due to multiple mechanisms, may complicate the course of patients with LOF FLNA variants and may be the presenting symptom leading to diagnosis. The combination of PH with parenchymal involvement and extrapulmonary symptoms (epilepsy, congenital heart diseases, valvular and aortic involvement, thrombocytopenia) should prompt genetic screening for FLNA.

A bovine model of hypoxia-induced pulmonary hypertension reveals a gradient of immune and matrisome response with a complement signature found in circulation.

Pulmonary hypertension (PH) is a progressive vascular disease characterized by vascular remodeling, stiffening, and luminal obstruction, driven by dysregulated cell proliferation, inflammation, and extracellular matrix (ECM) alterations. Despite the recognized contribution of ECM dysregulation to PH pathogenesis, the precise molecular alterations in the matrisome remain poorly understood. In this study, we employed a matrisome-focused proteomics approach to map the protein composition in a young bovine calf model of acute hypoxia-induced PH. Our findings reveal distinct alterations in the matrisome along the pulmonary vascular axis, with the most prominent changes observed in the main pulmonary artery. Key alterations included a strong immune response and wound repair signature, characterized by increased levels of complement components, coagulation cascade proteins, and provisional matrix markers. Additionally, we observed upregulation of ECM-modifying enzymes, growth factors, and core ECM proteins implicated in vascular stiffening, such as collagens, periostin, tenacsin-C, and fibrin(ogen). Notably, these alterations correlated with increased mean pulmonary arterial pressure and vascular remodeling. In the plasma, we identified increased levels of complement components, indicating a systemic inflammatory response accompanying the vascular remodeling. Our findings shed light on the dynamic matrisome remodeling in early-stage PH, implicating a wound-healing trajectory with distinct patterns from the MPA to the distal vasculature. This study provides novel insights into the molecular underpinnings of PH pathogenesis and highlights potential biomarkers and therapeutic targets within the matrisome landscape.

Super-Enhancer-Driven Syndecan-4 Regulates Intercellular Communication in Hypoxic Pulmonary Hypertension.

Unveiling pro-proliferation genes involved in crosstalk between pulmonary artery endothelial cells and pulmonary artery smooth muscle cells (PASMCs) are important to improving the therapeutic outcome of pulmonary hypertension (PH). Although growing studies have shown that super-enhancers (SEs) play a pivotal role in pathological and physiological processes, the SE-associated genes in PH and their impact on PASMC proliferation remain largely unexplored. We used serotype 5 adenovirus-associated virus to interfere with syndecan-4 and constructed an SU5416 combined with hypoxia-PH model. Chromatin immunoprecipitation sequencing analysis, chromatin immunoprecipitation quantitative polymerase chain reaction, and bioinformatics were used to confirm early growth response 1 was involved in regulating syndecan-4-associated SE in PASMCs. The effects of syndecan-4 and its underlying mechanisms were subsequently elucidated using Western blot, coimmunoprecipitation, and cell coculture assays. Herein, we identified a novel SE-associated gene, syndecan-4, in hypoxia-exposed PASMCs. Syndecan-4 was transcriptionally driven by early growth response 1 via an SE and was significantly overexpressed in hypoxic PASMCs and plasma from patients with PH. Mechanism studies revealed that syndecan-4 induces PASMC proliferation by interacting and regulating protein kinase C α ubiquitination. In addition, syndecan-4 was enriched in exosomes secreted from hypoxic PASMCs, which subsequently transported and led to pulmonary artery endothelial cell dysfunction. Syndecan-4 inhibition in hypoxia by serotype 5 adenovirus-associated virus treatment attenuated the pulmonary artery remodeling and development of PH invivo. Taken together, our results demonstrate that an SE-driven syndecan-4 modulates crosstalk of PASMCs and pulmonary artery endothelial cells and promotes vascular remodeling via the protein kinase C α and exosome pathway, thus providing potential targets for the early diagnosis and treatment of hypoxic PH.

Skeletal muscle-derived musclin attenuates glycolysis, oxidative stress, and pulmonary hypertension through the NPR3/AKT/mTORC1 pathway.

Exercise ameliorates pulmonary hypertension (PH) progression. However, the underlying mechanisms are largely unclear. Musclin is an exercise-responsive myokine that exerts protective effects on cardiovascular diseases. The current study aims to explore the role of musclin in the development of PH. A monocrotaline (MCT)-induced mouse PH model is established. Adeno-associated virus serotype 6 (AAV6)-mediated gene transfer is used to induce musclin overexpression in skeletal muscle. Ultrasound and morphological analyses are utilized to assess the severity of PH. Cell viability assay, Ki-67 immunofluorescence staining, wound healing assay, and transwell assay are used to evaluate the proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs). We find that the musclin levels in both plasma and skeletal muscle are decreased in MCT-treated mice. The external expression of musclin in skeletal muscle ameliorates pulmonary arterial remodeling and right ventricular dysfunction. In vitro, musclin treatment suppresses hypoxia-induced glycolysis, oxidative stress, proliferation, and migration. Further experiments reveal that musclin inhibits mechanistic target of rapamycin complex 1 (mTORC1) activity in hypoxia-stimulated PASMCs and pulmonary arteries of MCT-treated mice. Reactivating mTORC1 abolishes the protective role of musclin against PH. Additionally, musclin enhances its interaction with natriuretic peptide receptor 3 (NPR3) in PASMCs. Silencing of NPR3 reverses the inhibitory effects of musclin on AKT phosphorylation, mTORC1 activity, glycolysis, oxidative stress, proliferation, and migration in hypoxia-challenged PASMCs. In conclusion, our study highlights the inhibitory role of musclin in the proliferation and migration of PASMCs and PH progression, thereby providing a novel potent therapeutic strategy for treating PH and partly clarifying the mechanism of exercise-mediated protection against PH.

Suppression of Fibulin 5 Attenuated Pulmonary Arterial Hypertension Associated with Congenital Heart Disease via Mitigating Pulmonary Arterial Smooth Muscle Cell Dysfunction

Abstract Backgrounds: The specific molecular pathogenesis of pulmonary arterial hypertension associated with congenital heart disease (CHD-PAH) and the reversal still remain elusive. Fibulin 5 was involved in the regulation of cardiovascular system while little is known about the expression and functions of fibulin 5 in the development of CHD-PAH. This study aimed to investigate the role of fibulin 5 in the pathogenesis of the disease. Methods Lung samples were obtained both from patients with CHD-PAH and aortocaval shunt-associated PAH rat models. TGF-β1was used to induce human pulmonary artery smooth muscle cells (hPASMCs) dysfunction and lentiviruses were responsible for alteration of fibulin 5 expression. In an effort to explore the specific role of fibulin 5, an inhibitor of phosphatidylinositol-3-kinases (PI3K) /protein-serine-threonine kinase (AKT) which was defined as the TGF-β1-related downstream signaling was applied for further exploration. Results Fibulin 5 was pronouncedly elevated in the PASMCs of the remodeled pulmonary arterioles from patients with CHD-PAH and shunt-associated PAH rats. We found that over-expression of fibulin 5 could promote hPASMCs dysfunction induced by TGF-β1 and it could be reversed by the PI3K/AKT inhibitor LY294002 suggesting the essential role of TGF-β1/PI3K/AKT signaling activation in phenotypic switch, proliferation and migration of PASMCs. Moreover, an adeno-associated virus vector encoding fibulin 5 by intratracheally instillation with rats improve the hemodynamic parameters in shunt-associated PAH rats and its related pulmonary artery remodeling. Conclusion Over-expression of fibulin 5 could significantly promote hPASMCs dysfunction and pulmonary artery remodeling via reinforcing TGF-β1/PI3K/AKT signaling activation, which highlighted the potential valuable pharmacological target for the treatment of CHD-PAH.Fibulin 5 was over-expressed in PAHInhibition of fibulin 5 attenuated PAH

Ambrisentan reverses the prosenescent and anti-autophagic effect of high glucose on human pulmonary artery endothelial cells exposed to hypoxia

Abstract Background Cardiovascular (CV) comorbidities (systemic hypertension, hyperlipidemia, obesity, diabetes mellitus, peripheral arterial disease, coronary artery disease) are associated with reduced treatment response to specific drugs for group 1 pulmonary arterial hypertension (PAH). Often this epidemiological evidence is the result of misdiagnosis of group 2 PH associated with left heart disease, but it can also be explained as the intrinsic loss of the ability of pulmonary endothelial cells to respond to specific antiremodeling drugs. Aims To evaluate the impact of high glucose and hyperosmolar stress on the responsiveness of human pulmonary artery endothelial cells (hPAECs) to the endothelin receptor antagonist ambrisentan in terms of senescence, autophagic activity, viability and expression of some microRNAs involved in pulmonary arterial remodeling. Methods We incubated hPAECs with high glucose (HG, 30.5 mM) or high mannitol (HM, 25 mM), with/without ambrisentan (0.02 nM) in normoxia (N) or hypoxia (H) for 24 h and tested them for cell viability (MTT assay), protein and miRNA levels, autophagy and senescence (β-Gal assay) in vitro. Results H induced cell death as compared to N (752 ± 44 vs 701 ± 45 Abs units, p = 0.03). H reduced autophagy (Figure 1A-C) and induced senescence (Figure 1D), an effect further potentiated by HG and HM. Treatment with A in N and H significantly reversed the effect of HG but not HM on autophagy (Figure 1A-C) and senescence (Figure 1D). H increased the abundance of antiapoptotic miR124-3, compared to N in vehicle (V)-treated hPAEC (p=0.008) (Figure 2A), and induced a similar effect on antiapoptotic and proliferative miR191-3p, although not statistically significant (Figure 2B). In N, A potentiated the expression of both miR124-3p (p=0.007) and miR191-3p (p=0.02) in HG-treated hPAEC, and only miR191-3p in HM-treated hPAEC (p=0.01). A induced a similar effect on miR191-3p in hPAEC exposed to H+HG (p=0.02), with a non-statistically significant trend on miR124-3p. Conclusions In hPAEC exposed to H, A retains its pro-autophagic and antisenescent effects in an in vitro model mimicking diabetes. miR124-3p and miR191-3p may act as biomarkers of disease and treatment response to specific drugs in patients with PAH.Figure 1:autophagy and senescenceFigure 2:miRNA expression

PEDF upregulated in Ngfr-positive cells suppresses proliferation of pulmonary artery smooth muscle cells

Abstract Background Pulmonary arterial hypertension (PAH) is a disease with poor prognosis that causes right heart failure due to progressive pulmonary artery remodeling. Although existing pulmonary vasodilators contribute to pulmonary artery reverse remodeling by reducing share stress, the development of therapeutic agents that directly target pulmonary vascular remodeling is desired. Nerve growth factor receptor (Ngfr) is involved in the inflammatory reaction and repair process of damaged tissues. We have previously reported that Ngfr-positive cells are increased in the peripheral blood (PB) of PAH patients and are associated with disease progression. We also found that Ngfr inhibited the pathogenetic progression of pulmonary hypertension in a model of hypoxia-induced pulmonary hypertension. However, it remains unclear how Ngfr positive cells are involved in the pathology of PAH. Purpose In this study, we investigate how Ngfr-positive cells affect the pathogenesis of PAH. Methods & Results To evaluate the localization of Ngfr-positive cells in lung tissue, WT mice (C57BL/6) transplanted with bone marrow (BM) of GFP-Tg mice were kept under hypoxia for 3 weeks to create hypoxia-induced pulmonary hypertension (PH) model. Immunostaining of lung tissue sections indicated that double positive cells (Ngfr+GFP+) were present in the interstitial tissue around the small pulmonary artery. To examine the cellular characteristics of Ngfr-positive cells, RNA sequencing of Ngfr-positive and Ngfr-negative cells in PB was performed. Significantly 78 genes were upregulated in Ngfr-positive cells, including 11 secreted proteins. Among these, we focused on pigment epithelium-derived factor (PEDF), a secreted protein involved in angiogenesis. PEDF mRNA expression was decreased in the lung of Ngfr-KO mice compared to WT mice. We next examined the effect of PEDF on pulmonary artery smooth muscle cells (PASMCs). Platelet-derived growth factor (PDGF)-induced cell proliferation of PASMC was inhibited by PEDF stimulation. Furthermore, PEDF stimulation significantly inhibited PDGF-induced mRNA expressions of TNF and MMP9 in PASMCs. Conclusion In this study, we show that PEDF expression is upregulated in Ngfr-positive cells and that PEDF inhibits PDGF-induced hPASMC proliferation, suggesting that NGFR-positive cells may suppress pulmonary vascular remodeling through PEDF-mediated paracrine effects. PEDF might be a new therapeutic target for PH.

In vivo study of pulmonary artery remodeling and pulsatility in long Fontan patient

Abstract Background Pulmonary vasculopathy associated with Fontan circulation is a relatively unexplored entity that could have clinical and therapeutic implications. Purpose Our aim was to evaluate the physiology of the pulmonary artery in patients with Fontan circulation using conventional right heart catheterization (RHC) and intravascular ultrasound (IVUS). Methods From December 2022 to January 2024, data from all consecutive Fontan patients undergoing RHC were prospectively collected at a tertiary referral center for adult congenital heart disease. Pressures and pulmonary artery wall high-definition IVUS recordings (OptiCrossTM, 60 MHz, Boston Scientific) were recorded at rest and during apnea. Pathological Fontan pressure was defined when as exceeding 15 mmHg. IVUS pulsatility index was defined as [(systolic lumen area - diastolic lumen area)/(diastolic lumen area) ×100], Peterson's elastic modulus as (pulmonary pulse pressure/IVUS pulsatility index), and area of wall thickness as [(wall area - lumen area)/(lumen area) x100)]. In the literature, the mean ± standard deviation (SD) normal values of elastic modulus and wall thickness area have been reported as 21 ± 1.7 mmHg and 1.4 ± 1.3%, respectively, in healthy patients without pulmonary hypertension (1: doi: 10.1186/s12931-017-0568-z). We define an alteration in intrinsic pulmonary arterial wall viscoelastic properties as an elastic modulus > 70 mmHg, and we define structural pulmonary wall remodeling as an increase in wall thickness area greater than 5%. Results 13 patients were studied. Median age was 28.1 years interquartile range (IQR) (25.6- 41.4), 7 (53.9%) were female, and 4 (30,8%) were in functional New York Heart Associtation (NYHA) class I. RHC and pulmonary arterial wall IVUS values are presented in Figure 1. 6 (46.2%) had Fontan abnormal pressures at rest. All patients showed minimal pulse pressure in the pulmonary arteries synchronized with heart rate, median pulmonary pulse pressure (IQR) 3 (2-4) mmHg. This pulsatility was detected in the IVUS study in all patients, pulsatility index (median (IQR) 4.8 (3.3-10.1)) and elastic modulus (median (IQR) 51.5 (20.5-101.6) mmHg). All of them had some degree of structural pulmonary wall remodeling (>5% wall thickness area), and 10 (76.9%) of them had marked remodeling (> 10%). The median (IQR) of wall thickness area was 13.6 (10.5-18.3) % (Figure 2). We think that the pseudo-normality of the elastic modulus is due to a low pulmonary pulse pressure, however, structural remodeling could be secondary to the endothelial dysfunction present and described in arteries with very low pulsatility. Conclusions The IVUS evaluation of the pulmonary artery in Fontan patients may be useful for revealing significant information regarding pulmonary physiology. Pulmonary wall thickening has been observed, providing insight into altered structural remodeling of the pulmonary arteries in this population.

The causes of pulmonary hypertension and the benefits of aerobic exercise for pulmonary hypertension from an integrated perspective.

Pulmonary hypertension is a progressive disease of the pulmonary arteries that begins with increased pulmonary artery pressure, driven by progressive remodeling of the small pulmonary arteries, and ultimately leads to right heart failure and death. Vascular remodeling is the main pathological feature of pulmonary hypertension, but treatments for pulmonary hypertension are lacking. Determining the process of vascular proliferation and dysfunction may be a way to decipher the pathogenesis of pulmonary hypertension. In this review, we summarize the important pathways of pulmonary hypertension pathogenesis. We show how these processes are integrated and emphasize the benign role of aerobic exercise, which, as an adjunctive therapy, may be able to modify vascular remodeling in pulmonary hypertension.

Pulmonary Vascular Disease in Chronic Obstructive Pulmonary Disease.

Pulmonary hypertension (PH) is a vascular disease characterized by pulmonary artery remodeling and right heart failure. PH related to COPD is a precapillary form of the disease, with hemodynamic measurements including a mean pulmonary artery pressure of greater than 20 mm Hg, a wedge pressure of less than 15 mm Hg, and a pulmonary vascular resistance of greater than 3 WU (Woods units), categorized under the World Health Organization classification as group 3. The presence of PH in COPD has been known to increase morbidity and mortality. Limited studies have evaluated treatment options for PH related to COPD.

Sinomenine Hydrochloride Regulates the Apoptosis of Endothelial Cells through PPAR-γ to Alleviate PAH.

Pulmonary arterial hypertension is a progressive heart and lung disease that is caused by irreversible pulmonary vascular remodeling. Sinomenine hydrochloride is an alkaloid that is extracted from sinomenium acutum, which has strong anti-inflammatory effects. In this study, male rats were injected with monocrotaline, and endothelial cells were exposed to hypoxia for 24 hours to induce pulmonary arterial hypertension. Apoptosis, inflammation, and oxidative stress pathways were observed the in lungs and cells. Sinomenine hydrochloride repressed the increased right ventricular systolic pressure and attenuated the right ventricular hypertrophy and pulmonary artery remodeling in model rats. It reversed the expression of BCL2 and BAX and prevented the apoptosis of endothelial cells. Additionally, it increased the contents of IKBα and NRF2. P65, P-P65, TNFα, IL1β, and IL6 levels in the lungs decreased by it. Malondialdehyde contents decreased, and the superoxide dismutase and glutathione peroxidase activity and HO-1 level increased in the treatment group. In vivo, it promoted apoptosis of pulmonary artery endothelial cells. Moreover, by activating PPAR-γ, sinomenine hydrochloride attains the above effects. These data suggested that sinomenine hydrochloride could protect endothelial cells, restrain inflammation and oxidative stress, and enhance pulmonary vascular remodeling.

G6pdN126D Variant Increases the Risk of Developing VEGFR (Vascular Endothelial Growth Factor Receptor) Blocker-Induced Pulmonary Vascular Disease.

G6PD (glucose-6-phosphate-dehydrogenase) is a key enzyme in the glycolytic pathway and has been implicated in the pathogenesis of cancer and pulmonary hypertension-associated vascular remodeling. Here, we investigated the role of an X-linked G6pd mutation (N126D polymorphism), which is known to increase the risk of cardiovascular disease in individuals from sub-Saharan Africa and many others with African ancestry, in the pathogenesis of pulmonary hypertension induced by a vascular endothelial cell growth factor receptor blocker used for treating cancer. CRISPR-Cas9 genome editing was used to generate the G6pd variant (N126D; G6pdN126D) in rats. A single dose of the vascular endothelial cell growth factor receptor blocker sugen-5416 (SU; 20 mg/kg in DMSO), which is currently in a Phase 2/3 clinical trial for cancer treatment, was subcutaneously injected into G6pdN126D rats and their wild-type littermates. After 8 weeks of normoxic conditions, right ventricular pressure and hypertrophy, pulmonary artery remodeling, the metabolic profile, and cytokine expression were assessed. Right ventricular pressure and pulmonary arterial wall thickness were increased in G6PDN126D+SU/normoxic rats. Simultaneously, levels of oxidized glutathione, inositol triphosphate, and intracellular Ca2+ were increased in the lungs of G6PDN126D+SU/normoxic rats, whereas nitric oxide was decreased. Also increased in G6PDN126D+SU/normoxic rats were pulmonary levels of plasminogen activator inhibitor-1, thrombin-antithrombin complex, and expression of proinflammatory cytokines CCL3 (chemokine [C-C motif] ligand), CCL5, and CCL7. Our results suggest G6PDN126D increases inositol triphosphate-Ca2+ signaling, inflammation, thrombosis, and hypertrophic pulmonary artery remodeling in SU-treated rats. This suggests an increased risk of vascular endothelial cell growth factor receptor blocker-induced pulmonary hypertension in those carrying this G6PD variant.

Empagliflozin Attenuates Pulmonary Arterial Remodeling Through Peroxisome Proliferator-Activated Receptor Gamma Activation.

The loss of peroxisome proliferator-activated receptor gamma (PPARγ) exacerbates pulmonary arterial hypertension (PAH), while its upregulation reduces cell proliferation and vascular remodeling, thereby decreasing PAH severity. SGLT2 inhibitors, developed for type 2 diabetes, might also affect signal transduction in addition to modulating sodium-glucose cotransporters. Pulmonary arterial smooth muscle cells (PASMCs) isolated from patients with idiopathic pulmonary arterial hypertension (IPAH) were treated with three SGLT2 inhibitors, canagliflozin (Cana), dapagliflozin (Dapa), and empagliflozin (Empa), to investigate their antiproliferative effects. To assess the impact of Empa on PPARγ, luciferase reporter assays and siRNA-mediated PPARγ knockdown were employed to examine regulation of the γ-secretase complex and its downstream target Notch3. Therapy involving daily administration of Empa was initiated 21 days after inducing hypoxia-induced PAH in mice. Empa exhibited significant antiproliferative effects on fast-growing IPAH PASMCs. Empa activated PPARγ to prevent formation of the γ-secretase complex, with specific impacts on presenilin enhancer 2 (PEN2), which plays a crucial role in maintaining γ-secretase complex stability, thereby inhibiting Notch3. Similar results were obtained in lung tissue of chronically hypoxic mice. Empa attenuated pulmonary arterial remodeling and right ventricle hypertrophy in a hypoxic PAH mouse model. Moreover, PPARγ expression was significantly decreased and PEN2, and Notch3 levels were increased in lung tissue from PAH patients compared with non-PAH lung tissue. Empa reverses vascular remodeling by activating PPARγ to suppress the γ-secretase-Notch3 axis. We propose Empa as a PPARγ activator and potential therapeutic for PAH.

β-sitosterol alleviates pulmonary arterial hypertension by altering smooth muscle cell phenotype and DNA damage/cGAS/STING signaling

BackgroundPulmonary arterial smooth muscle cells (PASMCs) have a neoplastic phenotype characterized by hyperproliferative and anti-apoptotic features that contribute to pulmonary hypertension (PH) development. DNA-sensing adapter protein stimulator of interferon genes (STING) regulate the phenotypic switch of vessel smooth muscle cells. β-sitosterol (SITO) is a nutrient derived from plants that inhibits vascular smooth muscle cell proliferation without notable toxicity. However, the effect of SITO on cancer-like PH-associated pulmonary vascular remodeling and the specific mechanism has not yet be studied. PurposeThis study investigated the in vitro and in vivo effects of SITO against PH, and its underlying mechanisms. MethodsThe therapeutic efficacy of SITO was assessed, and its underlying mechanisms were explored in hypoxia-induced and platelet-derived growth factor (PDGF)-BB-stimulated primary PASMCs and in a monocrotaline (MCT)-induced preclinical PH rat model. SITO or sildenafil (SID) were administered after the MCT intraperitoneal injection. Pulmonary parameters, right heart function, morphology, and PASMCs were cultured for verification. The expression levels of DNA damage/cyclic GMP–AMP synthase (cGAS)/STING were determined using immunofluorescence and Western blotting. STING agonists that interfere with PASMCs were used to determine whether STING mediates the effects of SITO. ResultsSITO prevented PASMCs proliferation, promoted apoptosis and suppressed phenotypic switching in a dose-dependent manner in vitro and in vivo. In vivo results in rats demonstrated that four weeks of intragastric SITO administration effectively mitigated the MCT-induced elevation of hemodynamic parameters, improved right cardiac function, and reduced pulmonary arteries remodeling. Mechanistically, DNA damage and cGAS/STING/nuclear factor kappa-B signaling activation were observed in rats with PH and cultured PASMCs. SITO exhibited protective effects by suppressing the DNA damage, potentially via inhibiting the expression level of the cGAS/STING signaling pathway. Pharmacological overexpression of STING abolished the anti-proliferative effects of SITO treatment in hypoxia-induced and PDGF-stimulated PASMCs by downregulating PCNA. ConclusionSITO may be an attractive agent for PH vascular remodeling by inhibiting proliferation and modulating the phenotypic switch in PASMCs via the DNA damage/cGAS/STING signaling pathway. This study provides a novel therapeutic agent and mediator of the pathological development of PASMCs and PH.

TGFβ1, SMAD and β-catenin in pulmonary arteries of smokers, patients with small airway disease and COPD: potential drivers of EndMT.

We previously reported pulmonary arterial remodelling and active endothelial-to-mesenchymal transition (EndMT) in smokers and patients with early chronic obstructive pulmonary disease (COPD). In the present study, we aimed to evaluate the role of different drivers of EndMT. Immunohistochemical staining for EndMT drivers, TGF-β1, pSMAD-2/3, SMAD-7, and β-catenin, was performed on lung resections from 46 subjects. Twelve were non-smoker-controls (NC), six normal lung function smokers (NLFS), nine patients with small-airway diseases (SAD), nine mild-moderate COPD-current smokers (COPD-CS) and ten COPD-ex-smokers (COPD-ES). Histopathological measurements were done using Image ProPlus softwarev7.0. We observed lower levels of total TGF-β1 (P<0.05) in all smoking groups than in the non-smoking control (NC). Across arterial sizes, smoking groups exhibited significantly higher (P<0.05) total and individual layer pSMAD-2/3 and SMAD-7 than in the NC group. The ratio of SAMD-7 to pSMAD-2/3 was higher in COPD patients compared with NC. Total β-catenin expression was significantly higher in smoking groups across arterial sizes (P<0.05), except for COPD-ES and NLFS groups in small and medium arteries, respectively. Increased total β-catenin was positively correlated with total S100A4 in small and medium arteries (r = 0.35, 0.50; P=0.02, 0.01, respectively), with Vimentin in medium arteries (r = 0.42, P=0.07), and with arterial thickness of medium and large arteries (r = 0.34, 0.41, P=0.02, 0.01, respectively). This is the first study uncovering active endothelial SMAD pathway independent of TGF-β1 in smokers, SAD, and COPD patients. Increased expression of β-catenin indicates its potential interaction with SMAD pathway, warranting further research to identify the deviation of this classical pathway.

Unraveling the Mfn2-Warburg effect nexus: a therapeutic strategy to combat pulmonary arterial hypertension arising from catch-up growth after IUGR

BackgroundThe interplay between intrauterine and early postnatal environments has been associated with an increased risk of cardiovascular diseases in adulthood, including pulmonary arterial hypertension (PAH). While emerging evidence highlights the crucial role of mitochondrial pathology in PAH, the specific mechanisms driving fetal-originated PAH remain elusive.Methods and resultsTo elucidate the role of mitochondrial dynamics in the pathogenesis of fetal-originated PAH, we established a rat model of postnatal catch-up growth following intrauterine growth restriction (IUGR) to induce pulmonary arterial hypertension (PAH). RNA-seq analysis of pulmonary artery samples from the rats revealed dysregulated mitochondrial metabolic genes and pathways associated with increased pulmonary arterial pressure and pulmonary arterial remodeling in the RC group (postnatal catch-up growth following IUGR). In vitro experiments using pulmonary arterial smooth muscle cells (PASMCs) from the RC group demonstrated elevated proliferation, migration, and impaired mitochondrial functions. Notably, reduced expression of Mitofusion 2 (Mfn2), a mitochondrial outer membrane protein involved in mitochondrial fusion, was observed in the RC group. Reconstitution of Mfn2 resulted in enhanced mitochondrial fusion and improved mitochondrial functions in PASMCs of RC group, effectively reversing the Warburg effect. Importantly, Mfn2 reconstitution alleviated the PAH phenotype in the RC group rats.ConclusionsImbalanced mitochondrial dynamics, characterized by reduced Mfn2 expression, plays a critical role in the development of fetal-originated PAH following postnatal catch-up growth after IUGR. Mfn2 emerges as a promising therapeutic strategy for managing IUGR-catch-up growth induced PAH.