Artery Smooth Muscle Cell Proliferation Research Articles

Canagliflozin alleviates pulmonary hypertension by activating PPARγ and inhibiting its S225 phosphorylation.

Pulmonary hypertension (PH) is a progressive fatal disease with no cure. Canagliflozin (CANA), a novel medication for diabetes, has been found to have remarkable cardiovascular benefits. However, few studies have addressed the effect and pharmacological mechanism of CANA in the treatment of PH. Therefore, our study aimed to investigate the effect and pharmacological mechanism of CANA in treating PH. First, CANA suppressed increased pulmonary artery pressure, right ventricular hypertrophy, and vascular remodeling in both mouse and rat PH models. Network pharmacology, transcriptomics, and biological results suggested that CANA could ameliorate PH by suppressing excessive oxidative stress and pulmonary artery smooth muscle cell proliferation partially through the activation of PPARγ. Further studies demonstrated that CANA inhibited phosphorylation of PPARγ at Ser225 (a novel serine phosphorylation site in PPARγ), thereby promoting the nuclear translocation of PPARγ and increasing its ability to resist oxidative stress and proliferation. Taken together, our study not only highlighted the potential pharmacological effect of CANA on PH but also revealed that CANA-induced inhibition of PPARγ Ser225 phosphorylation increases its capacity to counteract oxidative stress and inhibits proliferation. These findings may stimulate further research and encourage future clinical trials exploring the therapeutic potential of CANA in PH treatment.

Sodium Houttuyfonate Alleviates MCT-induced Pulmonary Hypertension by Regulating Orai1 and Orai2.

Elevated intracellular Ca2+ concentration ([Ca2+]i) is a key trigger for pulmonary arterial smooth muscle cell (PASMC) proliferation and contributes greatly to pulmonary hypertension (PH). Extracellular Ca2+ influx via a store-operated Ca2+ channel (SOCC), termed store-operated Ca2+ entry (SOCE), is a crucial mechanism for [Ca2+]i elevation in PASMCs. Calcium release-activated calcium modulator (Orai) proteins, consisting of three members (Orai1-3), are the main components of SOCC. Sodium houttuyfonate (SH) is a product of the addition reaction of sodium bisulfite and houttuynin and has antibacterial, anti-inflammatory, and other properties. In this study, we assessed the contributions of Orai proteins to MCT-enhanced SOCE, [Ca2+]i, and cell proliferation in PASMCs and determined the effect of SH on MCT-PH and the underlying mechanism, focusing on Orai proteins, SOCE, and [Ca2+]i in PASMCs. Our results showed that 1) Orai1 and Orai2 were selectively upregulated in the distal pulmonary arteries (PAs) and the PASMCs of MCT-PH rats. 2) Knockdown of Orai1 or Orai2 reduced SOCE, [Ca2+]i, and cell proliferation without affecting their expression in PASMCs in MCT-PH rats. 3) SH significantly normalized the characteristic parameters in a dose-dependent manner in the MCT-PH rat model. 4) SH decreased MCT-enhanced SOCE, [Ca2+]i and PASMC proliferation via Orai1 or Orai2. These results indicate that SH likely exerts its protective role in MCT-PH by inhibiting the Orai1,2-SOCE-[Ca2+]i signaling pathway.

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.

MiR-221-3p is upregulated in acute pulmonary embolism complicated with pulmonary hypertension and promotes pulmonary arterial smooth muscle cells proliferation and migration by inhibiting PTEN

miR-221-3p is upregulated in acute pulmonary embolism complicated with pulmonary hypertension and promotes pulmonary arterial smooth muscle cells proliferation and migration by inhibiting PTEN

Curcumol reduces lower limb arteriosclerosis in rats by inhibiting human arterial smooth muscle cell activity.

Cardiovascular diseases, particularly those involving arterial stenosis and smooth muscle cell proliferation, pose significant health risks. This study aimed to investigate the therapeutic potential of curcumol in inhibiting platelet-derived growth factor-BB (PDGF-BB)-induced human aortic smooth muscle cell (HASMC) proliferation, migration and autophagy. Using cell viability assays, 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays and Western Blot analyses, we observed that curcumol effectively attenuated PDGF-BB-induced HASMC proliferation and migration in a concentration-dependent manner. Furthermore, curcumol mitigated PDGF-BB-induced autophagy, as evidenced by the downregulation of LC3-II/LC3-I ratio and upregulation of P62. In vivo experiments using an arteriosclerosis obliterans model demonstrated that curcumol treatment significantly ameliorated arterial morphology and reduced stenosis. Additionally, curcumol inhibited the activity of the KLF5/COX2 axis, a key pathway in vascular diseases. These findings suggest that curcumol has the potential to serve as a multi-target therapeutic agent for vascular diseases.

Hydroxycitric Acid Tripotassium Hydrate Attenuates Monocrotaline and Hypoxia-Induced Pulmonary Hypertension in Rats.

Pulmonary hypertension was induced in male Sprague-Dawley rats by a single subcutaneous injection of monocrotaline or hypoxic chamber. In vivo, inflammatory cytokine (including TNF-α, IL-1β, IL-6, and TGF-β, the level of SOD) expression, superoxide dismutase and hydrogen peroxide levels, and p-IκBα and p65 expressions were detected. In vitro, pulmonary artery smooth muscle cell proliferation and migration, ROS production, and hypoxia-inducible factor-1 expression were also studied. Hydroxycitric acid tripotassium hydrate decreased right ventricular systolic pressure and reduced right ventricular fibrosis and pulmonary vascular remodeling in rats with two kinds of pulmonary hypertension. Moreover, the expression of both inflammatory and oxidative stress factors was effectively reduced, and the p65 signaling pathway was found to be inhibited in this study. Additionally, hydroxycitric acid tripotassium hydrate inhibited human pulmonary artery smooth cell proliferation and migration in vitro. This study shows that hydroxycitric acid tripotassium hydrate can alleviate pulmonary hypertension caused by hypoxia and monocycloline in rats, improve remodeling of the right ventricle and pulmonary artery, and inhibit pulmonary artery smooth muscle cell proliferation and migration. The protective effects may be achieved by regulating inflammation and oxidative stress through the p65 signaling pathway.

Exposure to TCBPA stimulates the growth of arterial smooth muscle cells through the activation of the ROS/NF-κB/NLRP3 signaling pathway

Tetrachlorobisphenol A (TCBPA) and Tetrabromobisphenol S (TBBPS) are organic compounds widely used in industrial production, including in plastic and textile manufacturing. Presently, residual TCBPA is commonly detected in the environment as well as in human and animal sera. Therefore, it is imperative to assess the potential toxicological effects of TCBPA on organismal health. A series of biochemical experiments, including indirect immunofluorescence, ELISA, Western blot, MTT, etc, were conducted to analyze the effects of TCBPA on vascular smooth muscle cells. In this study, the biological impact of TCBPA on arterial smooth muscle cells (ASMCs) was investigated. CCK8 and EdU assays demonstrated significant proliferation of ASMCs following TCBPA treatment. Furthermore, TCBPA induced an inflammatory response in smooth muscle cells, as evidenced by the upregulated expression of inflammatory cytokines including IL-6, IL-1β, and MCP1. Additionally, we observed that TCBPA triggered an oxidative stress response in ASMCs by measuring ROS levels. To elucidate the underlying molecular mechanism of TCBPA-induced ASMC proliferation, we found that NLRP3 was essential for this process. Further investigation revealed that NLRP3 activation was mediated by NF-κB (which was activated by ROS). In summary, our findings suggest that TCBPA promotes the proliferation of ASMCs through the ROS/NF-κB/NLRP3 signaling cascade. This work indicates that TCBPA may represent a potential risk factor for the development of atherosclerosis, highlighting the need for judicious control of TCBPA usage.

FAK mediates hypoxia-induced pulmonary artery smooth muscle cell proliferation by modulating mitochondrial transcription termination factor 1/cyclin D1.

This study aimed to investigate the mechanism of FAK-dependent hypoxia-induced proliferation on human pulmonary artery smooth muscle cells (HPASMCs). Primary HPASMCs were isolated and cultured invitro under normal and hypoxia conditions to assess cell proliferation with cell counting kit-8. FAK and mitochondrial transcription termination factor 1 (mTERF1) were silenced with siRNA, mRNA, and protein levels of FAK, mTERF1, and cyclin D1 were determined. HPASMC proliferation increased under hypoxia compared to normal conditions. Knocking down FAK or mTERF1 with siRNA led to decreased cell proliferation under both normal and hypoxia conditions. FAK knockdown led to the reduction of both mTERF1 and cyclin D1 expressions under the hypoxia conditions, whereas mTERF1 knockdown led to the downregulation of cyclin D1 expression but not FAK expression under the same condition. However, under normal conditions, knocking down either FAK or mTERF1 had no impact on cyclin D1 expression. These results suggested that FAK may regulate the mTERF1/cyclin D1 signaling pathway to modulate cell proliferation in hypoxia.

EGR1 transcriptionally regulates SVEP1 to promote proliferation and migration in human coronary artery smooth muscle cells.

A low-frequency variant of sushi, von Willebrand factor type A, EGF, and pentraxin domain-containing protein 1 (SVEP1) is associated with the risk of coronary artery disease, as determined by a genome-wide association study. SVEP1 induces vascular smooth muscle cell proliferation and an inflammatory phenotype to promote atherosclerosis. In the present study, qRT‒PCR demonstrated that the mRNA expression of SVEP1 was significantly increased in atherosclerotic plaques compared to normal tissues. Bioinformatics revealed that EGR1 was a transcription factor for SVEP1. The results of the luciferase reporter assay, siRNA interference or overexpression assay, mutational analysis and ChIP confirmed that EGR1 positively regulated the transcriptional activity of SVEP1 by directly binding to its promoter. EGR1 promoted human coronary artery smooth muscle cell (HCASMC) proliferation and migration via SVEP1 in response to oxidized low-density lipoprotein (ox-LDL) treatment. Moreover, the expression level of EGR1 was increased in atherosclerotic plaques and showed a strong linear correlation with the expression of SVEP1. Our findings indicated that EGR1 binding to the promoter region drive SVEP1 transcription to promote HCASMC proliferation and migration.

A novel complement C3 inhibitor CP40-KK protects against experimental pulmonary arterial hypertension via an inflammasome NLRP3 associated pathway

BackgroundPulmonary arterial hypertension (PAH) is a severe cardiopulmonary disease characterized by complement dependent and proinflammatory activation of macrophages. However, effective treatment for complement activation in PAH is lacking. We aimed to explore the effect and mechanism of CP40-KK (a newly identified analog of selective complement C3 inhibitor CP40) in the PAH model.MethodsWe used western blotting, immunohistochemistry, and immunofluorescence staining of lung tissues from the monocrotaline (MCT)-induced rat PAH model to study macrophage infiltration, NLPR3 inflammasome activation, and proinflammatory cytokines (IL-1β and IL-18) release. Surface plasmon resonance (SPR), ELISA, and CH50 assays were used to test the affinity between CP40-KK and rat/human complement C3. CP40-KK group rats only received CP40-KK (2 mg/kg) by subcutaneous injection at day 15 to day 28 continuously.ResultsC3a was significantly upregulated in the plasma of MCT-treated rats. SPR, ELISA, and CH50 assays revealed that CP40-KK displayed similar affinity binding to human and rat complement C3. Pharmacological inhibition of complement C3 cleavage (CP40-KK) could ameliorate MCT-induced NLRP3 inflammasome activity, pulmonary vascular remodeling, and right ventricular hypertrophy. Mechanistically, increased proliferation of pulmonary arterial smooth muscle cells is closely associated with macrophage infiltration, NLPR3 inflammasome activation, and proinflammatory cytokines (IL-1β and IL-18) release. Besides, C3a enhanced IL-1β activity in macrophages and promoted pulmonary arterial smooth muscle cell proliferation in vitro.ConclusionOur findings suggest that CP40-KK treatment was protective in the MCT-induced rat PAH model, which might serve as a therapeutic option for PAH.

Blockade of ZFX Alleviates Hypoxia-Induced Pulmonary Vascular Remodeling by Regulating the YAP Signaling.

High expression of the zinc finger X-chromosomal protein (ZFX) correlates with proliferation, aggressiveness, and development in many types of cancers. In the current report, we investigated the efficacy of ZFX in mouse pulmonary artery smooth muscle cells (PASMCs) proliferation during pulmonary arterial hypertension (PAH). PASMCs were cultured in hypoxic conditions. Real-time PCR and western blotting were conducted to detect the expression of ZFX. Cell proliferation, apoptosis, migration, and invasion were, respectively, measured by CCK-8, flow cytometry, wound scratchy, and transwell assays. Glycolytic ability was validated by the extracellular acidification rate and oxygen consumption rate. Transcriptome sequencing technology was used to explore the genes affected by ZFX knockdown. Luciferase and chromatin immunoprecipitation assays were utilized to verify the possible binding site of ZFX and YAP1. Mice were subjected to hypoxia for 21days to induce PAH. The right ventricular systolic pressure (RVSP) was measured and ratio of RV/LV + S was calculated. The results show that ZFX was increased in hypoxia-induced PASMCs and mice. ZFX knockdown inhibited the proliferation, migration, and invasion of PASMC. Using RNA sequencing, we identify glycolysis and YAP as a key signaling of ZFX. ZFX knockdown inhibited Glycolytic ability. ZFX strengthened the transcription activity of YAP1, thereby regulating the YAP signaling. YAP1 overexpression reversed the effect of ZFX knockdown on hypoxia-treated PASMCs. In conclusion, ZFX knockdown protected mice from hypoxia-induced PAH injury. ZFX knockdown dramatically reduced RVSP and RV/(LV + S) in hypoxia-treated mice.

SCUBE1 promotes pulmonary artery smooth muscle cell proliferation and migration in acute pulmonary embolism by modulating BMP7.

After an episode of acute pulmonary embolism (APE), activated platelets have the ability to release various bioactive factors that can stimulate both proliferation and migration of pulmonary artery smooth muscle cells (PASMCs). SCUBE1 has been previously reported to engage in platelet-platelet interactions, potentially contributing to the activation of platelets in early onset thrombi. The purpose of this study was to examine the alterations in SCUBE1 expression in PASMCs after APE, as well as understand the mechanism behind these changes. The platelet-rich plasma samples of both APE patients and healthy individuals were collected. A hyperproliferative model of PASMCs was established by using platelet-derived growth factor (PDGF) as a stimulator and various assays were used to investigate how SCUBE1-mediated BMP7 can regulate PDGF-induced PASMC proliferation and migration. Elevated level of SCUBE1 were observed in platelet-rich plasma from patients with APE and in PASMCs induced by PDGF. SCUBE1 interference ameliorated PDGF-driven cell proliferation and migration, and also downregulated PCNA expression. Additionally, mechanistic studies demonstrated that SCUBE1 could directly bind to bone morphogenetic protein 7 (BMP7) and enhance BMP7 expression, which completely abolished the impact of SCUBE1 silencing on proliferation and migration ability of PASMCs after PDGF treatment. In the PDGF-induced proliferation of PASMCs, the expression of SCUBE1 and BMP7 was upregulated. Silencing of SCUBE1 impeded PDGF-induced proliferation and migration of PASMCs by restraining BMP7.

Metallothionein 3 Potentiates Pulmonary Artery Smooth Muscle Cell Proliferation by Promoting Zinc-MTF1-ATG5 Axis-mediated Autophagosome Formation

Metallothionein 3 Potentiates Pulmonary Artery Smooth Muscle Cell Proliferation by Promoting Zinc-MTF1-ATG5 Axis-mediated Autophagosome Formation

Intra-Amniotic Sildenafil and Rosiglitazone Late in Gestation Ameliorate the Pulmonary Hypertension Phenotype in Congenital Diaphragmatic Hernia

BackgroundPulmonary hypertension remains difficult to manage in congenital diaphragmatic hernia (CDH). Prenatal therapy may ameliorate postnatal pulmonary hypertension. We hypothesized that intra-amniotic (IA) injection of either sildenafil, a phosphodiesterase 5 inhibitor, or rosiglitazone, a PPAR-γ agonist, or both late in gestation would decrease the detrimental pulmonary vascular remodeling seen in CDH and improve peripheral pulmonary blood flow. MethodsPregnant rats were gavaged with nitrogen on embryonic day (E) 9.5 to induce fetal CDH. Sildenafil and/or rosiglitazone were administered to each fetus via an intra-amniotic injection after laparotomy on the pregnant dam at E19.5, and fetuses delivered at E21.5. Efficacy measures were gross necropsy, histology, peripheral blood flow assessment using intra-cardiac injection of a vascular tracer after delivery, and protein expression analysis. ResultsIntra-amniotic injections did not affect fetal survival, the incidence of CDH, or lung weight-to-body weight ratio in CDH fetuses. IA sildenafil injection decreased pulmonary vascular muscularization, and rosiglitazone produced an increase in peripheral pulmonary blood flow distribution. The combination of sildenafil and rosiglitazone decreased pulmonary artery smooth muscle cell proliferation. These intra-amniotic treatments did not show any negative effects in either CDH fetuses or control fetuses. ConclusionIA injection of sildenafil and rosiglitazone late in gestation ameliorates the pulmonary hypertensive phenotype of CDH and may have utility in clinical translation. Level of EvidenceNot applicable.

FUNDC1/USP15/Drp1 ameliorated TNF-α-induced pulmonary artery endothelial cell proliferation by regulating mitochondrial dynamics.

Mitochondrial dysfunction in pulmonary artery endothelial cells (PAECs) is related to the pathogenesis of pulmonary hypertension (PH). The mitochondrial receptor protein FUN14 domain containing 1 (FUNDC1) was found to be involved in pulmonary artery smooth muscle cell proliferation in PH. However, its role in PAECs remains unclear. We investigated FUNDC1 expression in the pulmonary artery endothelium in both monocrotaline-induced animal models and TNF-α-stimulated cell models. Additionally, the effect of FUNDC1 on PAECs proliferation and its possible mechanism were also investigated. We observed decreased FUNDC1 protein levels in animals and in vitro in PAECs. FUNDC1 deficiency in PAECs upregulated the expression of the deubiquitination enzyme ubiquitin-specific peptidase 15 (USP15), enhanced dynamin-related protein1 (Drp1)-mediated mitochondrial division, and increased mitochondrial ROS levels via the deubiquitination of Drp1. Additionally, FUNDC1 deficiency increased aerobic glycolysis, the production of ATP and lactic acid, and glucose uptake. FUNDC1 overexpression inhibited PAECs proliferation. Moreover, FUNDC1 overexpression in combination with a mitochondrial division or aerobic glycolysis inhibitor enhanced its inhibitory effect on cell proliferation. Our study findings suggest that FUNDC1 deficiency induced by inflammation can promote PAECs proliferation by regulating mitochondrial dynamics and cell energy metabolism via the USP15/Drp1 pathway.

A small-diameter vascular graft promotes rapid and benign remodeling of the neointima through dual release of nitric oxide and hydrogen sulfide

Coronary artery bypass grafting often requires autologous veins as alternatives. The limited source and problem of over-expansion are major obstacles for vein grafts, which could be solved by small-diameter artificial vascular graft. However, their clinical application is limited by thrombosis and restenosis caused by incomplete endothelialization and abnormal smooth muscle cells (SMCs) proliferation. Nitric oxide (NO) and hydrogen sulfide (H2S) are crucial signaling molecules in the cardiovascular system, known to regulate endothelial cells(ECs) and SMCs proliferation, migration, reduce oxidative stress, and inhibit inflammation to prevent intimal hyperplasia. In this study, we developed a new method to prepare a keratin based H2S donor(KAT) complexed with copper ions and electrospun it with PCL to prepare PCL/KAT-Cu small-diameter tissue-engineered vascular graft, which is capable of dual release of NO and H2S. We investigated the effects of NO and H2S release on human umbilical vein endothelial cells (HUVECs) and human umbilical arterial smooth muscle cells(HUASMCs) proliferation and migration, evaluated the graft's selectivity for HUVECs, and assessed its effects on macrophage phenotypic transitions and its protective effect on HUVECs under reactive oxygen species (ROS) conditions. Additionally, we tested the graft's ability to rapidly endothelialize under shear force using an in vitro 3D-perfusion system. After one month of in situ transplantation in rat abdominal aorta, the PCL/KAT-Cu small-diameter tissue-engineered vascular grafts demonstrated satisfactory intimal remodeling, comparable to natural blood vessels. In conclusion, our study presents a promising new strategy for remodeling small-diameter vascular grafts.

Matrine alleviates hypoxia-induced inflammation and pulmonary vascular remodelling via RPS5/NF-κB signalling pathway.

Hypoxia-induced vasoconstriction and vascular remodelling are the main pathological features of hypoxic pulmonary arterial hypertension (HPAH), and inflammation is participated in the occurrence of pulmonary vascular remodelling (PVR). Matrine is an alkaloid with the effects of anti-inflammation, antifibrosis and antitumour. But, few studies have explored the role of matrine in regulating PVR, and the related mechanisms are still unknown. In this study, we found that hypoxia-induced pulmonary artery smooth muscle cells(PASMCs) proliferation and inhibited its apoptosis, reduced the expression of ribosomal protein s5 and activated the nuclear factor kappa-B (NF-κB) signalling. Matrine, sildenafil and NF-κB inhibitor Bay 11-7082 could reverse these changes and impel the cell cycle in phase S retardation, and reduced the expression of p50, p65, proliferating cell nuclear antigen (PCNA), Bcl-2. In addition, matrine could lower right ventricular systolic pressure and mean pulmonary artery pressureof rats,α-smooth muscle actinand PCNA expression in pulmonary artery media, the levels of tumor necrosis factor-α and interleuki-1β, thus improved hypoxia-induced PVR. This study indicated that matrine could alleviate inflammation and improve PVR through reversing the imbalance of proliferation and apoptosis of PASMCs, thus it had a therapeutic effect on HPAH.

Canagliflozin ameliorates hypobaric hypoxia-induced pulmonary arterial hypertension by inhibiting pulmonary arterial smooth muscle cell proliferation

ABSTRACT Pulmonary arterial hypertension (PAH) is a disease with a high mortality and few treatment options to prevent the development of pulmonary vessel remodeling, pulmonary vascular resistance, and right ventricular failure. Canagliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor, is originally used in diabetes patients which could assist the glucose excretion and decrease blood glucose. Recently, a few studies have reported the protective effect of SGLT2 inhibitor on monocrotaline-induced PAH. However, the effects of canagliflozin on hypobaric hypoxia-induced PAH as well as its mechanism still unclear. In this study, we used hypobaric hypoxia-induced PAH mice model to demonstrate if canagliflozin could alleviate PAH and prevent pulmonary vessel remodeling. We found that daily canagliflozin administration significantly improved survival in mice with hypobaric hypoxia-induced PAH compared to vehicle control. Canagliflozin treatment significantly reduced right ventricular systolic pressure and increased pulmonary acceleration time determined by hemodynamic assessments. Canagliflozin significantly reduced medial wall thickening and decreased muscularization of pulmonary arterioles compared to vehicle treated mice. In addition, canagliflozin inhibited the proliferation and migration of pulmonary arterial smooth muscle cells through suppressing glycolysis and reactivating AMP-activated protein kinase signaling pathway under hypoxia condition. In summary, our findings suggest that canagliflozin is sufficient to inhibit pulmonary arterial remodeling which is a potential therapeutic strategy for PAH treatment.

Abstract 18691: Asporin Attenuates Pulmonary Arterial Smooth Muscle Cell Proliferation and Regulates Pulmonary Hypertension Development

Introduction: Pulmonary arterial hypertension (PAH) is a devastating pulmonary vascular lung disease characterized by increased pressure in the pulmonary arteries (PA) leading to cardiac right ventricular (RV) hypertrophy, RV failure, and ultimately death. Current PAH therapies merely improve symptoms temporarily and patients often need lung and heart transplantation. There is dire need for new insights into PAH pathophysiology to find novel therapeutic approaches. A key mechanism driving PAH is medial hyperplasia resulting from enhanced pulmonary arterial smooth muscle cell (PASMC) proliferation. Methods: We employed an unbiased bioinformatics approach in online-available microarray expression and single-cell sequencing datasets of dissected PA from PAH patients. In vitro experiments were performed in PASMC isolated from a female donor and PAH patient. Human lung tissue sections were obtained from the Pulmonary Hypertension Breakthrough Initiative Biobank. The Sugen/Hypoxia model was used to induce PH in rats. Results: We discovered that a novel gene, small leucine-rich proteoglycan Asporin (Aspn), is highly and significantly upregulated in PAH compared to donor PA and is most highly expressed in PASMC. We validated higher Aspn mRNA expression in human lungs and PASMC from PAH patients compared to controls, supporting our bioinformatics approach. Moreover, Aspn protein expression is elevated in PASMC in lung tissue sections from PAH patients compared to donors, as well as in PAH patient plasma . Functionally, silencing Aspn in PASMC from PAH patients promoted proliferation, whereas recombinant Aspn exerted opposite effects. To find potential downstream mediators of Aspn, STRING database analysis revealed that Aspn binds to the pro-proliferative mediator TGFβ1. Accordingly, TGFβ1 downstream mediator p-SMAD2/3 is enhanced upon Aspn silencing in PASMC. Intratracheal delivery of Aspn siRNA in rats aggravated Sugen/Hypoxia-induced PH, as assessed by RVSP, echocardiography and vascular remodeling. Conclusions: We hypothesize that elevated Aspn expression in PAH is a compensatory mechanism that inhibits PASMC proliferation, leading to decreased PA vascular remodeling, and that Aspn may be a novel therapeutic target against PAH.

GPR146 regulates pulmonary vascular remodeling by promoting pulmonary artery smooth muscle cell proliferation through 5-lipoxygenase

The pathological feature of hypoxic pulmonary hypertension (PH) is pulmonary vascular remodeling (PVR), primarily attributed to the hyperproliferation and apoptosis resistance of pulmonary artery smooth muscle cells (PASMCs). Existing PH-targeted drugs have difficulties in reversing PVR. Therefore, it is vital to discover a new regulatory mechanism for PVR and develop new targeted drugs. G protein-coupled receptor 146 (GPR146) is believed to participate in this process. This study aimed to investigate the role of GPR146 in PASMCs during PH. We investigated the role of GPR146 in PVR and its underlying mechanism using hypoxic PASMCs and mouse model (Sugen 5416 (20 mg/kg)/hypoxia). In our recent study, we have observed a significant increase in the expression of GPR146 protein in animal models of PH as well as in patients diagnosed with pulmonary arterial hypertension (PAH). Through immunohistochemistry, we found that GPR146 was mainly localized in the smooth muscle and endothelial layers of the pulmonary vasculature. GPR146 deficiency induction exhibited protective effects against hypoxia-induced elevation of right ventricular systolic blood pressure (RVSP), right ventricular hypertrophy, and pulmonary vascular remodeling in mice. In particular, the deletion of GPR146 attenuated the hypoxia-triggered proliferation of PASMCs. Furthermore, 5-lipoxygenase (5-LO) was related to PH development. Hypoxia and overexpression of GPR146 increased 5-LO expression, which was reversed through GPR146 knockdown or siRNA intervention. Our study discovered that GPR146 exhibited high expression in the pulmonary vessels of pulmonary hypertension. Subsequent research revealed that GPR146 played a crucial role in the development of hypoxic PH by promoting lipid peroxidation and 5-LO expression. In conclusion, GPR146 may regulate pulmonary vascular remodeling by promoting PASMCs proliferation through 5-LO, which presents a feasible target for PH prevention and treatment.