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.

CGRP attenuates pulmonary vascular remodeling by inhibiting the cGAS-STING-NFκB pathway in pulmonary arterial hypertension.

BACKGROUND: Hyperproliferation, inflammation, and mitochondrial abnormalities in pulmonary artery smooth muscle cells (PASMCs) underlie the pathological mechanisms of vascular remodeling in pulmonary arterial hypertension (PAH). Cytoplasmic mtDNA activates the cGAS-STING-NFκB pathway and secretes pro-inflammatory cytokines that may be involved in the pathogenesis of PAH. Calcitonin gene-related peptide (CGRP) acts as a vasodilator to regulate patterns of cellular energy metabolism and has vasodilatory and anti-inflammatory effects. METHODS: The role of the cGAS-STING-NFκB signaling pathway in PAH vascular remodeling and the regulation of CGRP in the cGAS-STING-NFκB signaling pathway were investigated by echocardiography, morphology, histology, enzyme immunoassay, and fluorometry. RESULTS: Monocrotaline (MCT) could promote right heart hypertrophy, pulmonary artery intima thickening, and inflammatory cell infiltration in rats. Cinnamaldehyde (CA)-induced CGRP release alleviates MCT-induced vascular remodeling in PAH. CGRP reduces PDGF-BB-induced proliferation, and migration, and downregulates smooth muscle cell phenotypic proteins. In vivo and in vitro experiments confirm that the mitochondria of PASMCs were damaged during PAH, and the superoxide and mtDNA produced by injured mitochondria activate the cGAS-STING-NFκB pathway to promote PAH process, while CGRP could play an anti-PAH role by protecting the mitochondria and inhibiting the cGAS-STING-NFκB pathway through PKA. CONCLUSION: This study identifies that CGRP attenuates cGAS-STING-NFκB axis-mediated vascular remodeling in PAH through PKA.

The siRNA-mediated knockdown of AP-1 restores the function of the pulmonary artery and the right ventricle by reducing perivascular and interstitial fibrosis and key molecular players in cardiopulmonary disease.

BACKGROUND: Pulmonary hypertension (PH) is a complex multifactorial vascular pathology characterized by an increased pulmonary arterial pressure, vasoconstriction, remodelling of the pulmonary vasculature, thrombosis in situ and inflammation associated with right-side heart failure. Herein, we explored the potential beneficial effects of treatment with siRNA AP-1 on pulmonary arterial hypertension (PAH), right ventricular dysfunction along with perivascular and interstitial fibrosis in pulmonary artery-PA, right ventricle-RV and lung in an experimental animal model of monocrotaline (MCT)-induced PAH. METHODS: Golden Syrian hamsters were divided into: (1) C group-healthy animals taken as control; (2) MCT group obtained by a single subcutaneous injection of 60 mg/kg MCT at the beginning of the experiment; (3) MCT-siRNA AP-1 group received a one-time subcutaneous dose of MCT and subcutaneous injections containing 100 nM siRNA AP-1, every two weeks. All animal groups received water and standard chow ad libitum for 12 weeks. RESULTS: In comparison with the MCT group, siRNA AP-1 treatment had significant beneficial effects on investigated tissues contributing to: (1) a reduction in TGF-ß1/ET-1/IL-1ß/TNF-α plasma concentrations; (2) a reduced level of cytosolic ROS production in PA, RV and lung and notable improvements regarding the ultrastructure of these tissues; a decrease of inflammatory and fibrotic marker expressions in PA (COL1A/Fibronectin/Vimentin/α-SMA/CTGF/Calponin/MMP-9), RV and lung (COL1A/CTGF/Fibronectin/α-SMA/F-actin/OB-cadherin) and an increase of endothelial marker expressions (CD31/VE-cadherin) in PA; (4) structural and functional recoveries of the PA [reduced Vel, restored vascular reactivity (NA contraction, ACh relaxation)] and RV (enlarged internal cavity diameter in diastole, increased TAPSE and PRVOFs) associated with a decrease in systolic and diastolic blood pressure, and heart rate; (5) a reduced protein expression profile of AP-1S3/ pFAK/FAK/pERK/ERK and a significant decrease in the expression levels of miRNA-145, miRNA-210, miRNA-21, and miRNA-214 along with an increase of miRNA-124 and miRNA-204. CONCLUSIONS: The siRNA AP-1-based therapy led to an improvement of pulmonary arterial and right ventricular function accompanied by a regression of perivascular and interstitial fibrosis in PA, RV and lung and a down-regulation of key inflammatory and fibrotic markers in MCT-treated hamsters.

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.

Effect of Benidipine Alone and in Combination With Bosentan and Sildenafil in Amelioration of Pulmonary Arterial Hypertension in Experimental Model in Rats.

ABSTRACT: Pulmonary arterial hypertension (PAH) is a persistent condition affecting the pulmonary arteries' endothelium. Benidipine, a calcium channel blocker, possesses vasodilatory, anti-inflammatory activity, reduces oxidative stress, and inhibits the activity of Transforming growth factor-ß (TGF-ß) and α-smooth muscle actin (α-SMA). The present study was designed to investigate the effect of benidipine alone and in combination with bosentan and sildenafil on monocrotaline (MCT)-induced pulmonary hypertension in a rat model. PAH was induced by a single-dose administration of MCT in rats. Animals were randomized into different groups and treated with benidipine alone and in combination with bosentan or sildenafil. Various parameters such as hemodynamic parameters, Fulton's index and oxidative stress parameters were performed. Additionally, histopathology of lung and right ventricular of heart tissue, immunohistochemistry, expression of α-SMA, endothelial nitric oxide synthase (eNOS), TGF-ß, and RT-PCR, and an in vitro study using human umbilical vein endothelial cells (HUVECs) was also carried out. Treatment of benidipine and its combination exhibited better prevention in the elevated right ventricular systolic pressure, right ventricular hypertrophy, rise in oxidative stress, and increase in expression of α-SMA and TGF-ß receptor 1 compared with MCT control group rats. In HUVECs, the expression of α-SMA was increased, whereas that of eNOS decreased after TGF-ß exposure and was substantially reversed after pretreatment with benidipine. We concluded that benidipine and its combination with bosentan and sildenafil exhibit beneficial effects in MCT-induced PAH through the eNOS/TGF-ß/α-SMA signaling pathway.

Mediastinal Cryobiopsy for Pathological Diagnosis of Fibrosing Mediastinitis-Associated Pulmonary Hypertension.

INTRODUCTION: Fibrosing mediastinitis is a benign but fatal disorder characterized by the proliferation of fibrous tissue in the mediastinum, causing encasement of mediastinal organs and extrinsic compression of adjacent bronchovascular structures. FM-associated pulmonary hypertension (FM-PH) is a serious complication of FM, resulting from the external compression of lung vessels. Pathologic assessment is important for etiologic diagnosis and effective treatment of this disease. CASE PRESENTATION: A 59-year-old male patient presented at our hospital and was diagnosed with FM-PH. He declined surgical biopsy that is the reference standard for pathologic assessment, in consideration of the potential risks. Therefore, an endobronchial ultrasound examination was performed, which identified the subcarinal lesion. Under ultrasound guidance, four needle aspirations were carried out, followed by one cryobiopsy. Histopathological examination of transbronchial needle aspiration specimens was inconclusive, while samples from cryobiopsy suggested a diagnosis of idiopathic FM. Further immunophenotyping demonstrated the infiltration of lymphocytes, macrophages, and FOXP3-positive cells in FM-PH. CONCLUSION: Mediastinal cryobiopsy might be a novel and safe option for FM-PH patients who are unwilling or unsuitable for surgical procedure.

The novel roles of YULINK in the migration, proliferation and glycolysis of pulmonary arterial smooth muscle cells: implications for pulmonary arterial hypertension.

BACKGROUND: Abnormal remodeling of the pulmonary vasculature, characterized by the proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs) along with dysregulated glycolysis, is a pathognomonic feature of pulmonary arterial hypertension (PAH). YULINK (MIOS, Entrez Gene: 54468), a newly identified gene, has been recently shown to possess pleiotropic physiologic functions. This study aims to determine novel roles of YULINK in the regulation of PAH-related pathogenesis, including PASMC migration, proliferation and glycolysis. RESULTS: Our results utilized two PAH-related cell models: PASMCs treated with platelet-derived growth factor (PDGF) and PASMCs harvested from monocrotaline (MCT)-induced PAH rats (PAH-PASMCs). YULINK modulation, either by knockdown or overexpression, was found to influence PASMC migration and proliferation in both models. Additionally, YULINK was implicated in glycolytic processes, impacting glucose uptake, glucose transporter 1 (GLUT1) expression, hexokinase II (HK-2) expression, and pyruvate production in PASMCs. Notably, YULINK and GLUT1 were observed to colocalize on PASMC membranes under PAH-related pathogenic conditions. Indeed, increased YULINK expression was also detected in the pulmonary artery of human PAH specimen. Furthermore, YULINK inhibition led to the suppression of platelet-derived growth factor receptor (PDGFR) and the phosphorylation of focal adhesion kinase (FAK), phosphoinositide 3-kinase (PI3K), and protein kinase B (AKT) in both cell models. These findings suggest that the effects of YULINK are potentially mediated through the PI3K-AKT signaling pathway. CONCLUSIONS: Our findings indicate that YULINK appears to play a crucial role in the migration, proliferation, and glycolysis in PASMCs and therefore positioning it as a novel promising therapeutic target for PAH.

Transcriptional profiles of pulmonary artery endothelial cells in pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is characterized by endothelial cell (EC) dysfunction. There are no data from living patients to inform whether differential gene expression of pulmonary artery ECs (PAECs) can discern disease subtypes, progression and pathogenesis. We aimed to further validate our previously described method to propagate ECs from right heart catheter (RHC) balloon tips and to perform additional PAEC phenotyping. We performed bulk RNA sequencing of PAECs from RHC balloons. Using unsupervised dimensionality reduction and clustering we compared transcriptional signatures from PAH to controls and other forms of pulmonary hypertension. Select PAEC samples underwent single cell and population growth characterization and anoikis quantification. Fifty-four specimens were analyzed from 49 subjects. The transcriptome appeared stable over limited passages. Six genes involved in sex steroid signaling, metabolism, and oncogenesis were significantly upregulated in PAH subjects as compared to controls. Genes regulating BMP and Wnt signaling, oxidative stress and cellular metabolism were differentially expressed in PAH subjects. Changes in gene expression tracked with clinical events in PAH subjects with serial samples over time. Functional assays demonstrated enhanced replication competency and anoikis resistance. Our findings recapitulate fundamental biological processes of PAH and provide new evidence of a cancer-like phenotype in ECs from the central vasculature of PAH patients. This "cell biopsy" method may provide insight into patient and lung EC heterogeneity to advance precision medicine approaches in PAH.

Pinocembrin attenuates susceptibility to atrial fibrillation in rats with pulmonary arterial hypertension.

BACKGROUND: Pulmonary arterial hypertension (PAH) is a disease characterized by pulmonary vascular remodeling that triggers fibrosis and excessive myocardium apoptosis, ultimately facilitating atrial fibrillation (AF). In various rat models, Pinocembrin has anti-fibrotic and anti-apoptotic effects, reducing arrhythmia vulnerability. However, whether pinocembrin alleviates to AF in a PAH model remains unclear. The experiment aims to investigate how pinocembrin affects AF susceptibility in PAH rats and the possible mechanisms involved. METHODS: The PAH model was induced by monocrotaline (MCT; i. p. 60 mg/kg). Concurrently, rats received pinocembrin (i.p.50 mg/kg) or saline. Hemodynamics parameters, electrocardiogram parameters, lung H.E. staining, atrial electrophysiological parameters, histology, Western blot, and TUNEL assay were detected. RESULTS: Compared to the control rats, MCT-induced PAH rats possessed prominently enhancive mPAP (mean pulmonary artery pressure), pulmonary vascular remodeling, AF inducibility, HRV, right atrial myocardial fibrosis, apoptosis, atrial ERP, APD, and P-wave duration. Additionally, there were lowered protein levels of Cav1.2, Kv4.2, Kv4.3, and connexin 40 (CX40) in the MCT group in right atrial tissue. However, pinocembrin reversed the above pathologies and alleviated the activity of the Rho A/ROCKs signaling pathway, including the expression of Rho A, ROCK1, ROCK2, and its downstream MYPT-1, LIMK2, BCL-2, BAX, cleaved-caspase3 in right atrial and HL-1 cells. CONCLUSION: Present data exhibited pinocembrin attenuated atrial electrical, ion-channel, and autonomic remodeling, diminished myocardial fibrosis and apoptosis levels, thereby reducing susceptibility to AF in the MCT-induced PAH rats. Furthermore, we found that pinocembrin exerted inhibitory action on the Rho A/ROCK signaling pathway, which may be potentially associated with its anti-AF effects.

Schistosomiasis-associated pulmonary hypertension unveils disrupted murine gut-lung microbiome and reduced endoprotective Caveolin-1/BMPR2 expression.

Schistosomiasis-associated Pulmonary Arterial Hypertension (Sch-PAH) is a life-threatening complication of chronic S. mansoni infection that can lead to heart failure and death. During PAH, the expansion of apoptosis-resistant endothelial cells (ECs) has been extensively reported; however, therapeutic approaches to prevent the progression or reversal of this pathological phenotype remain clinically challenging. Previously, we showed that depletion of the anti-apoptotic protein Caveolin-1 (Cav-1) by shedding extracellular vesicles contributes to shifting endoprotective bone morphogenetic protein receptor 2 (BMPR2) towards transforming growth factor beta (TGF-ß)-mediated survival of an abnormal EC phenotype. However, the mechanism underlying the reduced endoprotection in PAH remains unclear. Interestingly, recent findings indicate that, similar to the gut, healthy human lungs are populated by diverse microbiota, and their composition depends significantly on intrinsic and extrinsic host factors, including infection. Despite the current knowledge that the disruption of the gut microbiome contributes to the development of PAH, the role of the lung microbiome remains unclear. Thus, using a preclinical animal model of Sch-PAH, we tested whether S. mansoni infection alters the gut-lung microbiome composition and causes EC injury, initiating the expansion of an abnormal EC phenotype observed in PAH. Indeed, in vivo stimulation with S. mansoni eggs significantly altered the gut-lung microbiome profile, in addition to promoting injury to the lung vasculature, characterized by increased apoptotic markers and loss of endoprotective expression of lung Cav-1 and BMPR2. Moreover, S. mansoni egg stimulus induced severe pulmonary vascular remodeling, leading to elevated right ventricular systolic pressure and hypertrophy, characteristic of PAH. In vitro, exposure to the immunodominant S. mansoni egg antigen p40 activated TLR4/CD14-mediated transient phosphorylation of Cav-1 at Tyr14 in human lung microvascular EC (HMVEC-L), culminating in a mild reduction of Cav-1 expression, but failed to promote death and shedding of extracellular vesicles observed in vivo. Altogether, these data suggest that disruption of the host-associated gut-lung microbiota may be essential for the emergence and expansion of the abnormal lung endothelial phenotype observed in PAH, in addition to S. mansoni eggs and antigens.

Naked cuticle homolog 1 prevents mouse pulmonary arterial hypertension via inhibition of Wnt/ß-catenin and oxidative stress.

Pulmonary arterial hypertension (PAH) is a poorly prognostic cardiopulmonary disease characterized by abnormal contraction and remodeling of pulmonary artery (PA). Excessive proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs) are considered as the major etiology of PA remodeling. As a negative regulator of Wnt/ß-catenin pathway, naked cuticle homolog 1 (NKD1) is originally involved in the tumor growth and metastasis via affecting the proliferation and migration of different types of cancer cells. However, the effect of NKD1 on PAH development has not been investigated. In the current study, downregulated NKD1 was identified in hypoxia-challenged PASMCs. NKD1 overexpression by adenovirus carrying vector encoding Nkd1 (Ad-Nkd1) repressed hypoxia-induced proliferation and migration of PASMCs. Mechanistically, upregulating NKD1 inhibited excessive reactive oxygen species (ROS) generation and ß-catenin expression in PASMCs after hypoxia stimulus. Both inducing ROS and recovering ß-catenin expression abolished NKD1-mediated suppression of proliferation and migration in PASMCs. In vivo, we also observed decreased expression of NKD1 in dissected PAs of monocrotaline (MCT)-induced PAH model. Upregulating NKD1 by Ad-Nkd1 transfection attenuated the increase in right ventricular systolic pressure (RVSP), right ventricular hypertrophy index (RVHI), pulmonary vascular wall thickening, and vascular ß-catenin expression after MCT treatment. After recovering ß-catenin expression by SKL2001, the vascular protection of external expression of NKD1 was also abolished. Taken together, our data suggest that NKD1 inhibits the proliferation, migration of PASMC, and PAH via inhibition of ß-catenin and oxidative stress. Thus, targeting NKD1 may provide novel insights into the prevention and treatment of PAH.

Untangling the mechanisms of pulmonary arterial hypertension-induced right ventricular stiffening in a large animal model.

Pulmonary hypertension (PHT) is a devastating disease with low survival rates. In PHT, chronic pressure overload leads to right ventricle (RV) stiffening; thus, impeding diastolic filling. Multiple mechanisms may contribute to RV stiffening, including wall thickening, microstructural disorganization, and myocardial stiffening. The relative importance of each mechanism is unclear. Our objective is to use a large animal model to untangle these mechanisms. Thus, we induced pulmonary arterial hypertension (PAH) in sheep via pulmonary artery banding. After eight weeks, the hearts underwent anatomic and diffusion tensor MRI to characterize wall thickening and microstructural disorganization. Additionally, myocardial samples underwent histological and gene expression analyses to quantify compositional changes and mechanical testing to quantify myocardial stiffening. Finally, we used finite element modeling to disentangle the relative importance of each stiffening mechanism. We found that the RVs of PAH animals thickened most at the base and the free wall and that PAH induced excessive collagen synthesis, increased cardiomyocyte cross-sectional area, and led to microstructural disorganization, consistent with increased expression of fibrotic genes. We also found that the myocardium itself stiffened significantly. Importantly, myocardial stiffening correlated significantly with collagen synthesis. Finally, our computational models predicted that myocardial stiffness contributes to RV stiffening significantly more than other mechanisms. Thus, myocardial stiffening may be the most important predictor for PAH progression. Given the correlation between myocardial stiffness and collagen synthesis, collagen-sensitive imaging modalities may be useful for estimating myocardial stiffness and predicting PAH outcomes. STATEMENT OF SIGNIFICANCE: Ventricular stiffening is a significant contributor to pulmonary hypertension-induced right heart failure. However, the mechanisms that lead to ventricular stiffening are not fully understood. The novelty of our work lies in answering this question through the use of a large animal model in combination with spatially- and directionally sensitive experimental techniques. We find that myocardial stiffness is the primary mechanism that leads to ventricular stiffening. Clinically, this knowledge may be used to improve diagnostic, prognostic, and therapeutic strategies for patients with pulmonary hypertension.

Administration of A. muciniphila ameliorates pulmonary arterial hypertension by targeting miR-208a-3p/NOVA1 axis.

Pulmonary arterial hypertension (PH) is a chronic disease induced by a progressive increase in pulmonary vascular resistance and failure of the right heart function. A number of studies show that the development of PH is closely related to the gut microbiota, and lung-gut axis might be a potential therapeutic target in the PH treatment. A. muciniphila has been reported to play a critical role in treating cardiovascular disorders. In this study we evaluated the therapeutic effects of A. muciniphila against hypoxia-induced PH and the underlying mechanisms. Mice were pretreated with A. muciniphila suspension (2 × 108 CFU in 200 µL sterile anaerobic PBS, i.g.) every day for 3 weeks, and then exposed to hypoxia (9% O2) for another 4 weeks to induce PH. We showed that A. muciniphila pretreatment significantly facilitated the restoration of the hemodynamics and structure of the cardiopulmonary system, reversed the pathological progression of hypoxia-induced PH. Moreover, A. muciniphila pretreatment significantly modulated the gut microbiota in hypoxia-induced PH mice. miRNA sequencing analysis reveals that miR-208a-3p, a commensal gut bacteria-regulated miRNA, was markedly downregulated in lung tissues exposed to hypoxia, which was restored by A. muciniphila pretreatment. We showed that transfection with miR-208a-3p mimic reversed hypoxia-induced abnormal proliferation of human pulmonary artery smooth muscle cells (hPASMCs) via regulating the cell cycle, whereas knockdown of miR-208a-3p abolished the beneficial effects of A. muciniphila pretreatment in hypoxia-induced PH mice. We demonstrated that miR-208a-3p bound to the 3'-untranslated region of NOVA1 mRNA; the expression of NOVA1 was upregulated in lung tissues exposed to hypoxia, which was reversed by A. muciniphila pretreatment. Furthermore, silencing of NOVA1 reversed hypoxia-induced abnormal proliferation of hPASMCs through cell cycle modulation. Our results demonstrate that A. muciniphila could modulate PH through the miR-208a-3p/NOVA1 axis, providing a new theoretical basis for PH treatment.

The soluble receptor for advanced glycation end products is potentially predictive of pulmonary arterial hypertension in systemic sclerosis.

Introduction: Pulmonary arterial hypertension (PAH) and interstitial lung disease (ILD) are the leading causes of death in systemic sclerosis (SSc). Until now, no prospective biomarker to predict new onset of SSc-ILD or SSc-PAH in patients with SSc has reached clinical application. In homeostasis, the receptor for advanced glycation end products (RAGE) is expressed in lung tissue and involved in cell-matrix adhesion, proliferation and migration of alveolar epithelial cells, and remodeling of the pulmonary vasculature. Several studies have shown that sRAGE levels in serum and pulmonary tissue vary according to the type of lung-related complication. Therefore, we investigated levels of soluble RAGE (sRAGE) and its ligand high mobility group box 1 (HMGB1) in SSc and their abilities to predict SSc-related pulmonary complications. Methods: One hundred eighty-eight SSc patients were followed retrospectively for the development of ILD, PAH, and mortality for 8 years. Levels of sRAGE and HMGB1 were measured in serum by ELISA. Kaplan-Meier survival curves were performed to predict lung events and mortality and event rates were compared with a log-rank test. Multiple linear regression analysis was performed to examine the association between sRAGE and important clinical determinants. Results: At baseline, levels of sRAGE were significantly higher in SSc-PAH-patients (median 4099.0 pg/ml [936.3-6365.3], p = 0.011) and lower in SSc-ILD-patients (735.0 pg/ml [IQR 525.5-1988.5], p = 0.001) compared to SSc patients without pulmonary involvement (1444.5 pg/ml [966.8-2276.0]). Levels of HMGB1 were not different between groups. After adjusting for age, gender, ILD, chronic obstructive pulmonary disease, anti-centromere antibodies, the presence of puffy fingers or sclerodactyly, use of immunosuppression, antifibrotic therapy, or glucocorticoids, and use of vasodilators, higher sRAGE levels remained independently associated with PAH. After a median follow-up of 50 months (25-81) of patients without pulmonary involvement, baseline sRAGE levels in the highest quartile were predictive of development of PAH (log-rank p = 0.01) and of PAH-related mortality (p = 0.001). Conclusions: High systemic sRAGE at baseline might be used as a prospective biomarker for patients with SSc at high risk to develop new onset of PAH. Moreover, high sRAGE levels could predict lower survival rates due to PAH in patients with SSc.

The Evolution of Pulmonary Artery Denervation for Treatment of Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is a progressive, life-limiting disease. Despite significant medical progress over the last three decades, the prognosis of PAH remains poor. PAH is associated with sympathetic nervous system over-stimulation and baroreceptor-mediated vasoconstriction, leading to pathologic pulmonary artery (PA) and right ventricular remodeling. PA denervation is a minimally-invasive intervention that ablates local sympathetic nerve fibers and baroreceptors to modulate pathologic vasoconstriction. Preliminary animal and clinical studies have shown improvements in short-term pulmonary hemodynamics and PA remodeling. However, future studies are needed to elucidate appropriate patient selection, timing of intervention, and long-term efficacy before integration into standard of care.

Zinc promotes cell proliferation via regulating metal-regulatory transcription factor 1 expression and transcriptional activity in pulmonary arterial hypertension.

Metal responsive transcription factor 1 (MTF-1) is a zinc-dependent transcription factor involved in the development of pulmonary arterial hypertension (PAH), which is a life-threatening disease characterized by elevated pulmonary artery pressure and pulmonary vascular remodeling. However, little is known about the role and regulatory signaling of MTF-1 in PAH. This study aimed to investigate the effect and mechanism of MTF-1 on the proliferation of pulmonary arterial smooth muscle cells (PASMCs). Several techniques including intracellular-free zinc detected by fluorescent indicator-fluozinc-3-AM, western blot, luciferase reporter, and cell proliferation assay were conducted to perform a comprehensive analysis of MTF-1 in proliferation of PASMCs in PAH. Increased cytosolic zinc was shown in monocrotaline (MCT)-PASMCs and ZnSO4-treated PASMCs, which led to overexpression and overactivation of MTF-1, followed by the up-regulation of placental growth factor (PlGF). Elevated MTF-1 and PlGF were observed in western blot, and high transcriptional activity of MTF-1 was confirmed by luciferase reporter in ZnSO4-treated cells. Further investigation of cell proliferation revealed a favorable impact of zinc ions on PASMCs proliferation, with the deletion of Mtf-1/Plgf attenuating ZnSO4-induced proliferation. Flow cytometry analysis showed that blockade of PKC signaling inhibited the cell cycle of MCT-PASMCs and ZnSO4-treated PASMCs. The Zinc/PKC/MTF-1/PlGF pathway is involved in the up-regulatory effect on the PASMCs proliferation in the process of PAH. This study provided novel insight into zinc homeostasis in the pathogenesis of PAHs, and the regulation of MTF-1 might be a potential target for therapeutic intervention in PAH.

Application of transcriptomics and proteomics in pulmonary arterial hypertension.

The onset and progression of pulmonary arterial hypertension (PAH), a malignant disease, are associated with environmental and epigenetic factors. Recent advancements in transcriptomics and proteomics technology have provided new insights into PAH and identified novel gene targets involved in the development of the disease. Transcriptomic analysis has led to the discovery of possible novel pathways, such as miR-483 targeting several PAH-related genes and a mechanistic link between the increase in HERV-K mRNA and protein. Proteomic analysis has revealed crucial details, including the loss of SIRT3 activity and the significance of the CLIC4/Arf6 pathway in PAH pathogenesis. Gene profiles and protein interaction networks of PAH have been analyzed, clarifying the roles of differentially expressed genes or proteins in the occurrence and development of PAH. This article discusses these recent advances.

Ligustrazine alleviates pulmonary arterial hypertension in rats by promoting the formation of myocardin transcription complex in the nucleus of pulmonary artery smooth muscle cells.

Pulmonary arterial hypertension (PAH) is a pathophysiological state of abnormally elevated pulmonary arterial pressure caused by drugs, inflammation, toxins, viruses, hypoxia, and other risk factors. We studied the therapeutic effect and target of tetramethylpyrazine (tetramethylpyrazine [TMP]; ligustrazine) in the treatment of PAH and we speculated that dramatic changes in myocardin levels can significantly affect the progression of PAH. In vivo, the results showed that administration of TMP significantly prolonged the survival of PAH rats by reducing the proliferative lesions, right ventricular systolic pressure (RVSP), mean pulmonary arterial pressure (mPAP), and the Fulton index in the heart and lung of PAH rats. In vitro, TMP can regulate the levels of smooth muscle protein 22-alpha (SM22-α), and myocardin as well as intracellular cytokines such as NO, transforming growth factor beta (TGF-ß), and connective tissue growth factor (CTGF) in a dose-dependent manner (25, 50, or 100 µM). Transfection of myocardin small interfering RNA (siRNA) aggravated the proliferation of pulmonary artery smooth muscle cells (PSMCs), and the regulatory effect of TMP on α-smooth muscle actin (α-SMA) and osteopontin (OPN) disappeared. The application of 10 nM estrogen receptor alpha (ERα) inhibitor MPP promoted the proliferation of PSMCs, but it does not affect the inhibition of TMP on PSMCs proliferation. Finally, we found that TMP promoted the nucleation of myocardin-related transcription factor-A (MRTF-A) and combined it with myocardin. In conclusion, TMP can inhibit the transformation of PSMCs from the contractile phenotype to the proliferative phenotype by promoting the formation of the nuclear (MRTF-A/myocardin) transcription complex to treat PAH.

GATA6 coordinates cross-talk between BMP10 and oxidative stress axis in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a life-threatening condition characterized by a progressive increase in pulmonary vascular resistance leading to right ventricular failure and often death. Here we report that deficiency of transcription factor GATA6 is a shared pathological feature of PA endothelial (PAEC) and smooth muscle cells (PASMC) in human PAH and experimental PH, which is responsible for maintenance of hyper-proliferative cellular phenotypes, pulmonary vascular remodeling and pulmonary hypertension. We further show that GATA6 acts as a transcription factor and direct positive regulator of anti-oxidant enzymes, and its deficiency in PAH/PH pulmonary vascular cells induces oxidative stress and mitochondrial dysfunction. We demonstrate that GATA6 is regulated by the BMP10/BMP receptors axis and its loss in PAECs and PASMC in PAH supports BMPR deficiency. In addition, we have established that GATA6-deficient PAEC, acting in a paracrine manner, increase proliferation and induce other pathological changes in PASMC, supporting the importance of GATA6 in pulmonary vascular cell communication. Treatment with dimethyl fumarate resolved oxidative stress and BMPR deficiency, reversed hemodynamic changes caused by endothelial Gata6 loss in mice, and inhibited proliferation and induced apoptosis in human PAH PASMC, strongly suggesting that targeting GATA6 deficiency may provide a therapeutic advance for patients with PAH.

Pulmonary artery smooth muscle cell phenotypic switching: A key event in the early stage of pulmonary artery hypertension.

Pulmonary arterial hypertension (PAH) is a currently incurable pulmonary vascular disease. Since current research on PAH is mainly aimed at the middle and late stages of disease progression, no satisfactory results have been achieved. This has led researchers to focus on the early stages of PAH. This review highlights for the first time a key event in the early stages of PAH progression, namely, the occurrence of pulmonary arterial smooth muscle cell (PASMC) phenotypic switching. Summarizing the related reports of phenotypic switching provides new perspectives and directions for the early pathological progression and treatment strategies for PAH.