Pulmonary Hypertension In Rats Research Articles

Sildenafil Effect on Atrial Natriuretic Peptide Level in Pulmonary Hypertensive Rats.

Pulmonary Hypertension (PH) leads to changes in pulmonary vascular architecture, hypertrophy of the right ventricle, and heart failure. Sildenafil is a drug that can modulate PH by inducing smooth muscle relaxation and vasodilation. To investigate the ability of sildenafil to alleviate the monocritaline (MCT)-induced PH in rats and to estimate the role and its effect on the atrial natriuretic peptide (ANP) levels. 28 adult male rats were divided randomly into four groups: Group A (control group; n=7). Group B (MCT-treated group; n=7) was given a single dose of MCT 60 mg/kg subcutaneously. Group C (The reversal group; n=7) received a single dose of MCT 60 mg/kg subcutaneously for three weeks and then sildenafil at 50 mg/kg/day, given daily for another three weeks. Group D (The prevention group; n=7) simultaneously received a single dose of MCT 60 mg/kg subcutaneously and sildenafil daily at 50 mg/kg for three weeks. The animals in the prevention group showed a significant decrease in ANP levels compared to the reversal and MCT-treated groups. This decrease was associated with a significant reduction in the Fulton index ratio in the prevention group compared to the reversal group. The nitric oxide levels were also significantly higher in the reversal group than in the control group. Preventive sildenafil treatment was associated with a significant decrease in ANP levels and reduced MCT-induced cardiac hypertrophy in rats.

Pulmo-protection of long-term swimming exercise via improving insulin sensitivity in monocrotaline-induced pulmonary hypertensive rats

Exercise has been recognized as an effective intervention in the treatment of pulmonary arterial hypertension (PAH), supported by numerous studies. However, the precise effects of exercise on pulmonary function remain to be fully elucidated. In this study, using a rat model of swimming exercise training and monocrotaline-induced PAH, we aimed to explore its impact on pulmonary morphology and function. Our investigations revealed that MCT-treated rats exhibited augmented mean pulmonary arterial pressure (MPAP) and pulmonary vascular remodeling, which can be attenuated by 4 weeks of swimming exercise training (60 min/day, 5 days/week). Notably, MCT-treated rats showed impaired pulmonary function, as manifested by decreased tidal volume and dynamic compliance, which were reversed by exercise training. Assessment of pulmonary substrate in PAH rats indicated a prominent pro-inflammatory substrate, evidenced by macrophage accumulation through quantitative immunohistological analysis of macrophage-like cell expression (CD68), and extracellular matrix remodeling, evaluated by Masson staining. Importantly, both the pro-inflammatory substrate and extracellular matrix remodeling were ameliorated by swimming exercise training. Additionally, serum biochemical analysis demonstrated elevated levels of low-density lipoprotein cholesterol and Apolipoprotein B following MCT treatment, which were reduced with exercise intervention. Moreover, exercise enhanced systemic insulin sensitivity in both MCT-treated and untreated rats. Notably, MCT and exercise treatment both decreased fasting blood glucose (FBG) levels in rats, whereas exercise training reinstated FBG levels to normal in MCT-treated rats. In summary, our study suggests that swimming exercise confers a pulmonary protective effect in MCT-induced PAH rats, highlighting the potential importance of exercise-based rehabilitation in the management of PAH.

3D Imaging Reveals Complex Microvascular Remodeling in the Right Ventricle in Pulmonary Hypertension.

Pathogenic concepts of right ventricular (RV) failure in pulmonary arterial hypertension focus on a critical loss of microvasculature. However, the methods underpinning prior studies did not take into account the 3-dimensional (3D) aspects of cardiac tissue, making accurate quantification difficult. We applied deep-tissue imaging to the pressure-overloaded RV to uncover the 3D properties of the microvascular network and determine whether deficient microvascular adaptation contributes to RV failure. Heart sections measuring 250-µm-thick were obtained from mice after pulmonary artery banding (PAB) or debanding PAB surgery and properties of the RV microvascular network were assessed using 3D imaging and quantification. Human heart tissues harvested at the time of transplantation from pulmonary arterial hypertension cases were compared with tissues from control cases with normal RV function. Longitudinal 3D assessment of PAB mouse hearts uncovered complex microvascular remodeling characterized by tortuous, shorter, thicker, highly branched vessels, and overall preserved microvascular density. This remodeling process was reversible in debanding PAB mice in which the RV function recovers over time. The remodeled microvasculature tightly wrapped around the hypertrophied cardiomyocytes to maintain a stable contact surface to cardiomyocytes as an adaptation to RV pressure overload, even in end-stage RV failure. However, microvasculature-cardiomyocyte contact was impaired in areas with interstitial fibrosis where cardiomyocytes displayed signs of hypoxia. Similar to PAB animals, microvascular density in the RV was preserved in patients with end-stage pulmonary arterial hypertension, and microvascular architectural changes appeared to vary by etiology, with patients with pulmonary veno-occlusive disease displaying a lack of microvascular complexity with uniformly short segments. 3D deep tissue imaging of the failing RV in PAB mice, pulmonary hypertension rats, and patients with pulmonary arterial hypertension reveals complex microvascular changes to preserve the microvascular density and maintain a stable microvascular-cardiomyocyte contact. Our studies provide a novel framework to understand microvascular adaptation in the pressure-overloaded RV that focuses on cell-cell interaction and goes beyond the concept of capillary rarefaction.

Tibetan Mesenchymal Stem Cell-derived Exosomes Alleviate Pulmonary Vascular Remodeling in Hypoxic Pulmonary Hypertension Rats.

Hypoxic pulmonary hypertension (HPH) is characterized by progressive pulmonary vasoconstriction, vascular remodeling, and right ventricular hypertrophy, causing right heart failure. This study aimed to investigate the therapeutic effects of exosomes from Tibetan umbilical cord mesenchymal stem cells on HPH via the TGF-β1/Smad2/3 pathway, comparing them with exosomes from Han Chinese individuals. An HPH rat model was established in vivo, and a hypoxia-induced injury in the rat pulmonary artery smooth muscle cells (rPASMCs) was simulated in vitro. Exosomes from human umbilical cord mesenchymal stem cells were administered to HPH model rats or added to cultured rPASMCs. The therapeutic effects of Tibetan-mesenchymal stem cell-derived exosomes (Tibetan-MSC-exo) and Han-mesenchymal stem cell-derived exosomes (Han-MSC-exo) on HPH were investigated through immunohistochemistry, Western blotting, EdU, and Transwell assays. The results showed that Tibetan-MSC-exo significantly attenuated pulmonary vascular remodeling and right ventricular hypertrophy in HPH rats compared with Han-MSC-exo. Tibetan-MSC-exo demonstrated better inhibition of hypoxia-induced rPASMCs proliferation and migration. Transcriptome sequencing revealed upregulated genes (Nbl1, Id2, Smad6, and Ltbp1) related to the TGFβ pathway. Nbl1 knockdown enhanced hypoxia-induced rPASMCs proliferation and migration, reversing Tibetan-MSC-exo-induced downregulation of TGFβ1 and p-Smad2/3. Furthermore, TGFβ1 overexpression hindered the therapeutic effects of Tibetan-MSC-exo and Han-MSC-exo on hypoxic injury. These findings suggest that Tibetan-MSC-exo favors HPH treatment better than Han-MSC-exo, possibly through the modulation of the TGFβ1/Smad2/3 pathway via Nbl1.

Berberine Alleviates Right Heart Dysfunction in MCT-Induced PAH Rats by Targeting Myocardial Mitochondrial Oxidative Phosphorylation

Background: Right ventricular failure (RVF) stands as a leading cause of mortality among pulmonary hypertension (PAH) patients. The underlying mechanisms contributing to Right Ventricular (RV) dysfunction remain enigmatic, with a conspicuous absence of effective treatments. RV functional deterioration is influenced by mitochondria oxidative phosphorylation and fatty acid oxidation. Berberine (BBR), a natural active compound extracted from Berberis vulgaris rhizomes, has shown promise for treating cardiovascular diseases due to its interactions with multiple intracellular targets. This study aimed to investigate the applicability and therapeutic effcacy of BBR in RVF and explore the mechanisms involved in its beneficial effects in monocrotaline (MCT)-induced PAH rats. Methods: Right ventricular hypertrophy (RVH) and RVF were induced via a single injection of MCT (60 mg/kg, ip) in Sprague-Dawley rats. BBR (4 mg/kg body weight per day) was administered through oral gavage. RVH and RVF were confirmed through transthoracic echocardiography (TTE). Transmission electron microscopy (TEM) was utilized to observe changes in cardiomyocyte and mitochondrial ultrastructure within the right ventricle tissue. Comparisons were made among Normal control (NC), RVH, RVH+BBR, RVF, and RVF+ BBR groups. RNA sequencing (RNA-seq) of right ventricle tissue was performed for both the NC (treated with normal saline, ip) and right ventricle dysfunction (RVH and RVF) groups. Gene Ontology and KEGG enrichment analyses were conducted using the R-package “Clusterprofiler.” RV transcriptome results were validated through qPCR and Western blot analysis. Results: Echocardiography and elevated ANP and BNP levels confirmed the presence of RVF following the development of MCT-induced PAH. TEM imaging revealed significant disruptions in mitochondrial ultrastructure, particularly in a subset of interfibrillar mitochondria, which exhibited swelling, disorganization, and reduced cristae density in RVH rats. These disruptions were even more pronounced in RVF rats. Transcriptomic profiles of RV tissue from RVH and RVF identified a total of 175 differentially expressed genes (DEGs). Gene Ontology and KEGG enrichment analyses pinpointed 38 genes associated with oxidative phosphorylation. qPCR demonstrated downregulation of mitochondrial oxidative phosphorylation-related genes (NDUFA2, NDUFA13, COX7B, COX6C, GSTK1, ECH1) in RVF. BBR ameliorated right heart function, leading to decreased RV internal diameter at end-diastole, a reduced RV hypertrophy index, and an increased RV fractional area change. This was accompanied by the recovery of ANP and BNP levels. Furthermore, after BBR application, the mitochondrial structure within RV tissue exhibited marked improvement. Lastly, Western blot analysis revealed that fatty acid oxidation-related proteins (Decr1, Acadm, Acacb, Hadh, Hadhb, Prkag3, Acaa2) were downregulated in right heart dysfunction, and BBR effectively mitigated the decline in oxidation-related protein levels in both RVH and RVF. Conclusion: In summary, our results underscore the critical role of BBR in preserving cardiac function from RVF in MCT-induced PAH rats. This preservation appears to occur through the upregulation of mitochondrial oxidative phosphorylation levels, ultimately enhancing energy metabolism within the mitochondria of RV cardiomyocytes in the right heart of PAH. This work is supported by Natural Science Foundation of Shandong Province(ZR2022MH052), China Postdoctoral Fund (NO.2021M691944) to XMY. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

MiR-210-5p Promotes Pulmonary Hypertension by Blocking ATP2A2.

Aberrant expression of ATPase sarcoplasmic/endoplasmic retic Ca2+ transporting 2 (ATP2A2) has attracted attention for its pathophysiologic role in pulmonary hypertension (PH). Several miRNAs, including miR-210-5p, have also been reported to be pathogenic factors in PH, but their exact mechanisms remain unknown. This study aimed to elucidate the potential mechanisms of miR-210-5p and ATP2A2 in MCT-induced PH. Eighteen Sprague-Dawley rats were randomly divided into two groups-monoclonal (MCT) group and control group-and then administered MCT (60mg/kg) and saline, respectively. mPAP, PVR, RVHI, WT%, and WA% were significantly increased in PH rats after 3weeks, confirming that the modeling of PH rats was successful. Subsequently, we determined the expression of ATP2A2 and miR-210-5p in lung tissues using WB and qRT-PCR methods. We established an in vitro model using BMP4 and TGF-β1 treatment of pulmonary artery smooth muscle cells (PASMCs) and examined the expression of ATP2A2 and miR-210-5p using the same method. To further elucidate the regulatory relationship between ATP2A2 and miR-210-5p, we altered the expression level of miR-210-5p and detected the corresponding changes in ATP2A2 levels. In addition, we demonstrated the relationship by dual luciferase experiments. Finally, the effect of silencing ATP2A2 could be confirmed by the level of cell membrane Ca2+ in PAMSCs. Up-regulation of miR-210-5p and down-regulation of ATP2A2 were observed in the MCT group compared with the control group, which was confirmed in the in vitro model. In addition, elevated miR-210-5p expression decreased the level of ATP2A2 while increasing the proliferation of PASMCs, and the results of the dual luciferase assay further confirmed that ATP2A2 is a downstream target of miR-210-5p. Additionally, silencing ATP2A2 resulted in increased cytoplasmic Ca2+ levels in PAMSCs. In MCT-induced PH, miR-210-5p promotes pulmonary vascular remodeling by inhibiting ATP2A2.

TRPC4 aggravates hypoxic pulmonary hypertension by promoting pulmonary endothelial cell apoptosis

Pulmonary hypertension (PH) is a devastating disease that lacks effective treatment options and is characterized by severe pulmonary vascular remodeling. Pulmonary arterial endothelial cell (PAEC) dysfunction drives the initiation and pathogenesis of pulmonary arterial hypertension. Canonical transient receptor potential (TRPC) channels, a family of Ca2+-permeable channels, play an important role in various diseases. However, the effect and mechanism of TRPCs on PH development have not been fully elucidated. Among the TRPC family members, TRPC4 expression was markedly upregulated in PAECs from hypoxia combined with SU5416 (HySu)-induced PH mice and monocrotaline (MCT)-treated PH rats, as well as in hypoxia-exposed PAECs, suggesting that TRPC4 in PAECs may participate in the occurrence and development of PH. In this study, we aimed to investigate whether TRPC4 in PAECs has an aggravating effect on PH and elucidate the molecular mechanisms. We observed that hypoxia treatment promoted PAEC apoptosis through a caspase-12/endoplasmic reticulum stress (ERS)-dependent pathway. Knockdown of TRPC4 attenuated hypoxia-induced apoptosis and caspase-3/caspase-12 activity in PAECs. Accordingly, adeno-associated virus (AAV) serotype 6-mediated pulmonary endothelial TRPC4 silencing (AAV6-Tie-shRNA-TRPC4) or TRPC4 antagonist suppressed PH progression as evidenced by reduced right ventricular systolic pressure (RVSP), pulmonary vascular remodeling, PAEC apoptosis and reactive oxygen species (ROS) production. Mechanistically, unbiased RNA sequencing (RNA-seq) suggested that TRPC4 deficiency suppressed the expression of the proapoptotic protein sushi domain containing 2 (Susd2) in hypoxia-exposed mouse PAECs. Moreover, TRPC4 activated hypoxia-induced PAEC apoptosis by promoting Susd2 expression. Therefore, inhibiting TRPC4 ameliorated PAEC apoptosis and hypoxic PH in animals by repressing Susd2 signaling, which may serve as a therapeutic target for the management of PH.

Pulmonary hypertension impairs vasomotor function in rat diaphragm arterioles

Pulmonary hypertension (PH) is a chronic, progressive condition in which respiratory muscle dysfunction is a primary contributor to exercise intolerance and dyspnea in patients. Contractile function, blood flow distribution, and the hyperemic response are altered in the diaphragm with PH, and we sought to determine whether this may be attributed, in part, to impaired vasoreactivity of the resistance vasculature. We hypothesized that there would be blunted endothelium-dependent vasodilation and impaired myogenic responsiveness in arterioles from the diaphragm of PH rats. Female Sprague-Dawley rats were randomized into healthy control (HC, n = 9) and monocrotaline-induced PH rats (MCT, n = 9). Endothelium-dependent and -independent vasodilation and myogenic responses were assessed in first-order arterioles (1As) from the medial costal diaphragm in vitro. There was a significant reduction in endothelium-dependent (via acetylcholine; HC, 78 ± 15% vs. MCT, 47 ± 17%; P < 0.05) and -independent (via sodium nitroprusside; HC, 89 ± 10% vs. MCT, 66 ± 10%; P < 0.05) vasodilation in 1As from MCT rats. MCT-induced PH also diminished myogenic constriction (P < 0.05) but did not alter passive pressure responses. The diaphragmatic weakness, impaired hyperemia, and blood flow redistribution associated with PH may be due, in part, to diaphragm vascular dysfunction and thus compromised oxygen delivery which occurs through both endothelium-dependent and -independent mechanisms.

HMGB2 Release Promotes Pulmonary Hypertension and Predicts Severity and Mortality of Patients With Pulmonary Arterial Hypertension.

Pulmonary hypertension (PH) is a progressive and life-threatening disease characterized by pulmonary vascular remodeling, which involves aberrant proliferation and apoptosis resistance of the pulmonary arterial smooth muscle cells (PASMCs), resembling the hallmark characteristics of cancer. In cancer, the HMGB2 (high-mobility group box 2) protein promotes the pro-proliferative/antiapoptotic phenotype. However, the function of HMGB2 in PH remains uninvestigated. Smooth muscle cell (SMC)-specific HMGB2 knockout or HMGB2-OE (HMGB2 overexpression) mice and HMGB2 silenced rats were used to establish hypoxia+Su5416 (HySu)-induced PH mouse and monocrotaline-induced PH rat models, respectively. The effects of HMGB2 and its underlying mechanisms were subsequently elucidated using RNA-sequencing and cellular and molecular biology analyses. Serum HMGB2 levels were measured in the controls and patients with pulmonary arterial (PA) hypertension. HMGB2 expression was markedly increased in the PAs of patients with PA hypertension and PH rodent models and was predominantly localized in PASMCs. SMC-specific HMGB2 deficiency or silencing attenuated PH development and pulmonary vascular remodeling in hypoxia+Su5416-induced mice and monocrotaline-treated rats. SMC-specific HMGB2 overexpression aggravated hypoxia+Su5416-induced PH. HMGB2 knockdown inhibited PASMC proliferation in vitro in response to PDGF-BB (platelet-derived growth factor-BB). In contrast, HMGB2 protein stimulation caused the hyperproliferation of PASMCs. In addition, HMGB2 promoted PASMC proliferation and the development of PH by RAGE (receptor for advanced glycation end products)/FAK (focal adhesion kinase)-mediated Hippo/YAP (yes-associated protein) signaling suppression. Serum HMGB2 levels were significantly increased in patients with PA hypertension, and they correlated with disease severity, predicting worse survival. Our findings indicate that targeting HMGB2 might be a novel therapeutic strategy for treating PH. Serum HMGB2 levels could serve as a novel biomarker for diagnosing PA hypertension and determining its prognosis.

Reduced exercise capacity occurs before intrinsic skeletal muscle dysfunction in experimental rat models of pulmonary hypertension.

Reduced exercise capacity in pulmonary hypertension (PH) significantly impacts quality of life. However, the cause of reduced exercise capacity in PH remains unclear. The objective of this study was to investigate whether intrinsic skeletal muscle changes are causative in reduced exercise capacity in PH using preclinical PH rat models with different PH severity. PH was induced in adult Sprague-Dawley (SD) or Fischer (CDF) rats with one dose of SU5416 (20 mg/kg) injection, followed by 3 weeks of hypoxia and additional 0-4 weeks of normoxia exposure. Control s rats were injected with vehicle and housed in normoxia. Echocardiography was performed to assess cardiac function. Exercise capacity was assessed by VO2 max. Skeletal muscle structural changes (atrophy, fiber type switching, and capillary density), mitochondrial function, isometric force, and fatigue profile were assessed. In SD rats, right ventricular systolic dysfunction is associated with reduced exercise capacity in PH rats at 7-weektimepoint in comparison to control rats, while no changes were observed in skeletal muscle structure, mitochondrial function, isometric force, or fatigue profile. CDF rats at 4-weektimepoint developed a more severe PH and, in addition to right ventricular dysfunction, the reduced exercise capacity in these rats is associated with skeletal muscle atrophy; however, mitochondrial function, isometric force, and fatigue profile in skeletal muscle remain unchanged. Our data suggest that cardiopulmonary impairments in PH are the primary cause of reduced exercise capacity, which occurs before intrinsic skeletal muscle dysfunction.

Collagen triple helix repeat-containing protein 1 is a novel biomarker of right ventricular involvement in pulmonary hypertension

BackgroundPulmonary hypertension leads to right ventricular failure, which is a major determinant of prognosis. Circulating biomarkers for right ventricular function are poorly explored in pulmonary hypertension. This study aimed to clarify the significance of collagen triple helix repeat-containing protein 1 (CTHRC1) as a biomarker of right ventricular failure in pulmonary hypertension. MethodsA monocrotaline-induced pulmonary hypertension rat model was used to evaluate right ventricular CTHRC1 expression and its relationship with fibrosis. Next, human plasma CTHRC1 levels were measured in controls (n = 20), pulmonary arterial hypertension (n = 46), and chronic thromboembolic pulmonary hypertension (CTEPH) patients (n = 64) before the first and after the final balloon pulmonary angioplasty. ResultsCTHRC1 expression was higher in the right ventricles of rats with monocrotaline-induced pulmonary hypertension than in those of controls. CTHRC1 was co-localized with vimentin and associated with fibrosis in the right ventricles. Plasma CTHRC1 levels were higher in human pulmonary arterial hypertension (P = 0.006) and CTEPH patients (P = 0.011) than in controls. Plasma CTHRC levels were correlated with B-type natriuretic peptide (R = 0.355, P < 0.001), tricuspid lateral annular peak systolic velocity (R = -0.213, P = 0.029), and right ventricular fractional area change (R = -0.225, P = 0.017). Finally, plasma CTHRC1 levels were decreased after the final balloon pulmonary angioplasty (P < 0.001) in CTEPH. ConclusionsCTHRC1 can be a circulating biomarker associated with right ventricular function and fibrosis in pulmonary hypertension and might reflect the therapeutic efficacy of balloon pulmonary angioplasty in CTEPH.

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.

Retracted: Curcumin Improves Pulmonary Hypertension Rats by Regulating Mitochondrial Function.

[This retracts the article DOI: 10.1155/2021/1078019.].

Effect of hydrogen inhalation on cardiovascular and interstitial components of pulmonary hypertension in rats

Hydrogen is known to have selective antioxidant properties. It binds highly reactive hydroxyl radicals. The pathogenesis of the monocrotaline animal model of pulmonary hypertension is associated with oxidative stress and leads to all the symptoms of pulmonary hypertension (PH) and interstitial lung disease (ILD) associated with hypertension.The aim of this work was to study the effect of 4% hydrogen inhalations on the symptoms of PH and ILD in rats.Methods. To model monocrotaline-induced pulmonary hypertension (MCT-PH), two groups of animals received a single subcutaneous injection of monocrotaline (MCT) on day 1. The control group was injected subcutaneously with MCT solvent only. The animals receiving MCT were further divided into 2 subgroups. Subgroup 1 rats breathed room air and subgroup 2 rats breathed a mixture of room air and 4% hydrogen. The regular inhalations continued until day 21. On day 21, hemodynamic parameters were measured under urethane anesthesia, the heart and its components and the lungs were weighed, and lung tissue was preserved for morphological study.Results. The inhalation had no effect on the main cardiovascular symptoms of PH, but a positive effect on the state of the connective tissue of the lungs affected by PH was shown. The mast cell response was reduced both quantitatively and functionally. There was a decrease in tryptase expression by mast cells, with predominance of the forms without signs of degranulation. TGF-β secretion was also significantly reduced and visualized by immunopositive cells in alveolar cellular structures and vessel walls.Conclusion. Inhalation of 4% hydrogen reduces inflammation and fibrosis of lung tissue during the development of MCT-PH.

Ntsr1 contributes to pulmonary hypertension by enhancing endoplasmic reticulum stress via JAK2-STAT3-Thbs1 signaling

Pulmonary hypertension (PH) is a severe clinical syndrome with pulmonary vascular remodeling and poor long-term prognosis. Neurotensin receptor 1 (Ntsr1), serve as one of the G protein-coupled receptors (GPCRs), implicates in various biological processes, but the potential effects of Ntsr1 in PH development are unclear. The Sugen/Hypoxia (SuHx) or monocrotaline (MCT) induced rat PH model was used in our study and the PH rats showed aggravated pulmonary artery remodeling and increased right ventricular systolic pressure (RVSP). Our results revealed that Ntsr1 induced endoplasmic reticulum (ER) stress response via ATF6 activation contributed to the development of PH. Moreover, RNA-sequencing (RNA-seq) and phosphoproteomics were performed and the Ntsr1-JAK2-STAT3-thrombospondin 1 (Thbs1)-ATF6 signaling was distinguished as the key pathway. In vitro, pulmonary artery smooth muscle cells (PASMCs) under hypoxia condition showed enhanced proliferation and migration properties, which could be inhibited by Ntsr1 knockdown, JAK2 inhibitor (Fedratinib) treatment, STAT3 inhibitior (Stattic) treatment, Thbs1 knockdown or ATF6 knockdown. In addition, adeno-associated virus 1 (AAV1) were used to knockdown the expression of Ntsr1, Thbs1 or ATF6 in rats and reversed the phenotype of PH. In summary, our results reveal that Ntsr1-JAK2-STAT3-Thbs1 pathway can induce enhanced ER stress via ATF6 activation and increased PASMC proliferation and migration capacities, which can be mechanism of the pulmonary artery remodeling and PH. Targeting Ntsr1 might be a novel therapeutic strategy to ameliorate PH.

Up-regulated expression of two-pore domain K+ channels, KCNK1 and KCNK2, is involved in the proliferation and migration of pulmonary arterial smooth muscle cells in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a severe and rare disease in the cardiopulmonary system. Its pathogenesis involves vascular remodeling of the pulmonary artery, which results in progressive increases in pulmonary arterial pressure. Chronically increased pulmonary arterial pressure causes right ventricular hypertrophy and subsequent right heart failure. Pulmonary vascular remodeling is attributed to the excessive proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs), which are induced by enhanced Ca2+ signaling following the up-/down-regulation of ion channel expression. In the present study, the functional expression of two-pore domain potassium KCNK channels was investigated in PASMCs from idiopathic PAH (IPAH) patients and experimental pulmonary hypertensive (PH) animals. In IPAH-PASMCs, the expression of KCNK1/TWIK1 and KCNK2/TREK1 channels was up-regulated, whereas that of KCNK3/TASK1 and KCNK6/TWIK2 channels was down-regulated. The similar up-regulated expression of KCNK1 and KCNK2 channels was observed in the pulmonary arterial smooth muscles of monocrotaline-induced PH rats, Sugen 5416/hypoxia-induced PH rats, and hypoxia-induced PH mice. The facilitated proliferation of IPAH-PASMCs was suppressed by the KCNK channel blockers, quinine and tetrapentylammonium. The migration of IPAH-PASMCs was also suppressed by these channel blockers. Furthermore, increases in the proliferation and migration were inhibited by the siRNA knockdown of KCNK1 or KCNK2 channels. The siRNA knockdown also caused membrane depolarization and subsequent decrease in cytosolic [Ca2+]. The phosphorylated level of c-Jun N-terminal kinase (JNK) was elevated in IPAH-PASMCs compared to normal-PASMCs. The increased phosphorylation was significantly reduced by the siRNA knockdown of KCNK1 or KCNK2 channels. Collectively, these findings indicate that the up-regulated expression of KCNK1 and KCNK2 channels facilitates the proliferation and migration of PASMCs via enhanced Ca2+ signaling and JNK signaling pathway, which is associated with vascular remodeling in PAH.

Gut Fungal Microbiota Alterations in Pulmonary Arterial Hypertensive Rats.

The gut microbiome's imbalance has been implicated in the pathogenesis of pulmonary arterial hypertension (PAH), yet the contribution of the gut mycobiome remains largely unclear. This study delineates the gut mycobiome profile in PAH and examines its interplay with the bacterial microbiome alterations. Fecal samples from monocrotaline-induced PAH rats and matched controls were subjected to internal transcribed spacer 1 (ITS1) sequencing for fungal community assessment and 16S ribosomal RNA (rRNA) gene sequencing for bacterial community characterization. Comparative analysis revealed no significant disparities in the overall mycobiome diversity between the PAH and control groups. However, taxonomic profiling identified differential mycobiome compositions, with the PAH group exhibiting a significant enrichment of genera such as Wallemia, unidentified_Branch02, Postia, Malassezia, Epicoccum, Cercospora, and Alternaria. Conversely, genera Xeromyces, unidentified_Plectosphaerellaceae, and Monilia were more abundant in the controls. Correlations of Malassezia and Wallemia abundance with hemodynamic parameters were observed. Indications of bidirectional fungal-bacterial community interactions were also noted. This investigation reveals distinct gut mycobiome alterations in PAH, which are intricately associated with concurrent bacterial microbiome changes, suggesting a possible contributory role of gut fungi in PAH pathophysiology. These findings underscore the potential for novel gut mycobiome-targeted therapeutic interventions in PAH management.

Jagged 2 inhibition attenuates hypoxia-induced mitochondrial damage and pulmonary hypertension through Sirtuin 1 signaling.

Notch pathway has played a significant role in the pathophysiology of pulmonary hypertension (PH). However, the role of Jagged 2 (Jag2), one ligand of Notch, remains to be elucidated.Therefore, determining the contribution of Jag2 to PH and its impact on pulmonary artery smooth muscle cells (PASMCs) was the aim of this investigation. Adeno-associated virus-mediated Jag2 inhibition was used to explore the role of Jag2 in peripheral pulmonary vascular remodeling assessed in a rat model of chronic hypoxia (10% O2, 4 weeks) induced pulmonary hypertension. In vitro, the effect of Jag2 silencing on hypoxia (1% O2, 24h) induced rat PASMCs was determined. Group differences were assessed using a 2-sided unpaired Student's t-test for two groups and one-way ANOVA for multiple groups. Jag2 upregulation was first confirmed in rats with sustained hypoxia-induced PH using publicly available gene expression data, experimental PH rat models and hypoxia induced rat PASMCs. Jag2 deficiency decreased oxidative stress injury, peripheral pulmonary vascular remodeling (0.276±0.020 vs. 0.451±0.033 μm, P<0.001, <50μm), and right ventricular systolic pressure (36.8±3.033 vs. 51.8±4.245 mmHg, P<0.001) in the chronic hypoxia-induced rat model of PH. Moreover, Jag2 knockdown decreased proliferation (1.227±0.051 vs. 1.45±0.07, P = 0.012), increased apoptosis (16.733%±0.724% vs. 6.56%±0.668%, P<0.001), and suppressed mitochondrial injury in hypoxia-treated rat PASMCs. Jag2 inhibition restored the activity of the Nrf2/HO-1 pathway, which was abolished by Sirtuin 1 deficiency. These findings show that Jag2 is essential for modulating pulmonary vascular dysfunction and accelerating PH, and that inhibition of Jag2 expression suppresses the progression and development of PH.

Role of the microtubule network in the passive anisotropic viscoelasticity of right ventricle with pulmonary hypertension progression

Cardiomyocytes are viscoelastic and contribute significantly to right ventricle (RV) mechanics. Microtubule, a cytoskeletal protein, has been shown to regulate cardiomyocyte viscoelasticity. Additionally, hypertrophied cardiomyocytes from failing myocardium have increased microtubules and cell stiffness. How the microtubules contribute to the tissue-level viscoelastic behavior in RV failure remains unknown. Our aim was to investigate the role of the microtubules in the passive anisotropic viscoelasticity of the RV free wall (RVFW) during pulmonary hypertension (PH) progression. Equibiaxial stress relaxation tests were conducted in the RVFW from healthy and PH rats under early (6%) and end (15%) diastolic strains, and at sub- (1Hz) and physiological (5Hz) stretch-rates. The RVFW viscoelasticity was also measured before and after the depolymerization of microtubules at 5Hz. In intact tissues, PH increased RV viscosity and elasticity at both stretch rates and strain levels, and the increase was stronger in the circumferential than longitudinal direction. At 6% of strain, the removal of microtubules reduced elasticity, viscosity, and the ratio of viscosity to elasticity in both directions and for both healthy and diseased RVs. However, at 15% of strain, the effect of microtubules was different between groups – both viscosity and elasticity were reduced in healthy RVs, but in the diseased RVs only the circumferential viscosity and the ratio of viscosity to elasticity were reduced. These data suggest that, at a large strain with collagen recruitment, microtubules play more significant roles in healthy RV tissue elasticity and diseased RV tissue viscosity. Our findings suggest cardiomyocyte cytoskeletons are critical to RV passive viscoelasticity under pressure overload. Statement of SignificanceThis study investigated the impact of microtubules on the passive anisotropic viscoelasticity of the right ventricular (RV) free wall at healthy and pressure-overloaded states. We originally found that the microtubules contribute significantly to healthy and diseased RV viscoelasticity in both (longitudinal and circumferential) directions at early diastolic strains. At end diastolic strains (with the engagement of collagen fibers), microtubules contribute more to the tissue elasticity of healthy RVs and tissue viscosity of diseased RVs. Our findings reveal the critical role of microtubules in the anisotropic viscoelasticity of the RV tissue, and the altered contribution from healthy to diseased state suggests that therapies targeting microtubules may have potentials for RV failure patients.

Endothelial phosphodiesterase 4B inactivation ameliorates endothelial-to-mesenchymal transition and pulmonary hypertension

Pulmonary hypertension (PH) is a fatal disorder characterized by pulmonary vascular remodeling and obstruction. The phosphodiesterase 4 (PDE4) family hydrolyzes cyclic AMP (cAMP) and is comprised of four subtypes (PDE4A–D). Previous studies have shown the beneficial effects of pan-PDE4 inhibitors in rodent PH; however, this class of drugs is associated with side effects owing to the broad inhibition of all four PDE4 isozymes. Here, we demonstrate that PDE4B is the predominant PDE isozyme in lungs and that it was upregulated in rodent and human PH lung tissues. We also confirmed that PDE4B is mainly expressed in the lung endothelial cells (ECs). Evaluation of PH in Pde4b wild type and knockout mice confirmed that Pde4b is important for the vascular remodeling associated with PH. In vivo EC lineage tracing demonstrated that Pde4b induces PH development by driving endothelial-to-mesenchymal transition (EndMT), and mechanistic studies showed that Pde4b regulates EndMT by antagonizing the cAMP-dependent PKA–CREB–BMPRII axis. Finally, treating PH rats with a PDE4B-specific inhibitor validated that PDE4B inhibition has a significant pharmacological effect in the alleviation of PH. Collectively, our findings indicate a critical role for PDE4B in EndMT and PH, prompting further studies of PDE4B-specific inhibitors as a therapeutic strategy for PH.