Mechanisms underlying age-associated exacerbation of pulmonary veno-occlusive disease.

Pulmonary veno-occlusive disease (PVOD) is a rare but severe form of pulmonary hypertension characterized by the obstruction of pulmonary arteries and veins, causing increased pulmonary artery pressure and leading to right ventricular (RV) heart failure. PVOD is often resistant to conventional pulmonary arterial hypertension (PAH) treatments and has a poor prognosis, with a median survival time of 2-3 years after diagnosis. We previously showed that the administration of a chemotherapy agent mitomycin C (MMC) in rats mediates PVOD through the activation of the eukaryotic initiation factor 2 (eIF2) kinase protein kinase R (PKR) and the integrated stress response (ISR), resulting in the impairment of vascular endothelial junctional structure and barrier function. Here, we demonstrate that aged rats over 1 year exhibit more severe vascular remodeling and RV hypertrophy than young adult rats following MMC treatment. This is attributed to an age-associated elevation of basal ISR activity and depletion of protein phosphatase 1, leading to prolonged eIF2 phosphorylation and sustained ISR activation. Pharmacological blockade of PKR or ISR mitigates PVOD phenotypes in both age groups, suggesting that targeting the PKR/ISR axis could be a potential therapeutic strategy for PVOD.

Upregulation of Angiomotin-Like 2 Ameliorates Experimental Pulmonary Arterial Hypertension by Inactivating YAP1 Signaling.

ABSTRACT: Angiomotin-like 2 (AMOTL2) is related to numerous physiological and pathological conditions by affecting signal transduction. However, whether AMOTL2 is linked to pulmonary arterial hypertension (PAH) has not been addressed. This work aimed to investigate the potential role of AMOTL2 in PAH. A decrease in AMOTL2 abundance was observed in the lungs of PAH rats. The upregulation of AMOTL2 significantly decreased right ventricle systolic pressure and right ventricular hypertrophy in PAH rats. Overexpression of AMOTL2 also led to a noteworthy decrease in vascular wall thickness, pulmonary artery area, and collagen deposition in rats with PAH. AMOTL2 was downregulated in hypoxia-stimulated pulmonary arterial smooth muscle cells (PASMCs). Moreover, AMOTL2 overexpression impeded hypoxia-evoked proliferation, migration, and phenotypic transformation in rat PASMCs. Mechanistic investigation revealed that Yes-associated protein 1 (YAP1) activation in PAH rats or hypoxia-stimulated PASMCs was markedly inhibited by AMOTL2 overexpression, which was associated with increased large tumor suppressor 1/2 phosphorylation. The inhibition of large tumor suppressor 1/2 reversed the AMOTL2-mediated inactivation of YAP1. Restoring the activity of YAP1 reversed the inhibitory effect of AMOTL2 on hypoxia-evoked proliferation, migration, and phenotypic transformation of PASMCs. Collectively, these results suggest that AMOTL2 can ameliorate PAH in a rat model by interfering with pulmonary arterial remodeling via the inactivation of YAP1 signaling. Our work indicates that AMOTL2 may be a candidate target for novel drug development for the treatment of PAH.

PAI-1 deficiency drives pulmonary vascular smooth muscle remodeling and pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is a progressive disease characterized by vasoconstriction and remodeling of small pulmonary arteries (PAs). Central to the remodeling process is a switch of pulmonary vascular cells to a proliferative, apoptosis-resistant phenotype. Plasminogen activator inhibitors-1 and -2 (PAI-1 and PAI-2) are the primary physiological inhibitors of urokinase-type and tissue-type plasminogen activators (uPA and tPA), but their roles in PAH are unsettled. Here, we report that: 1) PAI-1, but not PAI-2, is deficient in remodeled small PAs and in early-passage PA smooth muscle and endothelial cells (PASMCs and PAECs) from subjects with PAH compared with controls; 2) PAI-1-/- mice spontaneously develop pulmonary vascular remodeling associated with upregulation of mTORC1 signaling, pulmonary hypertension (PH), and right ventricle (RV) hypertrophy; and 3) pharmacological inhibition of uPA in human PAH PASMCs suppresses proproliferative mTORC1 and SMAD3 signaling, restores PAI-1 levels, reduces proliferation, and induces apoptosis in vitro, and prevents the development of SU5416/hypoxia-induced PH and RV hypertrophy in vivo in mice. These data strongly suggest that downregulation of PAI-1 in small PAs promotes vascular remodeling and PH due to unopposed activation of uPA and consequent upregulation of mTOR and transforming growth factor-ß (TGF-ß) signaling in PASMCs, and call for further studies to determine the potential benefits of targeting the PAI-1/uPA imbalance to attenuate and/or reverse pulmonary vascular remodeling and PH.NEW & NOTEWORTHY This study identifies a novel role for the deficiency of plasminogen activator inhibitor (PAI)-1 and resultant unrestricted uPA activity in PASMC remodeling and PH in vitro and in vivo, provides novel mechanistic link from PAI-1 loss through uPA-induced Akt/mTOR and TGFß-Smad3 upregulation to pulmonary vascular remodeling in PH, and suggests that inhibition of uPA to rebalance the uPA-PAI-1 tandem might provide a novel approach to complement current therapies used to mitigate this pulmonary vascular disease.

Nur77 induced by HIF-1α mediates vascular remodeling in hypoxic pulmonary hypertension.

Nur77 is a member of the NR4A subfamily of orphan nuclear receptors that is expressed and has a function within the immune system. This study aimed to investigate the role of Nur77 in hypoxic pulmonary hypertension. SPF male SD rats were exposed in hypobaric chamber simulating 5000 m high altitude for 0, 3, 7, 14, 21 or 28 days. Rat pulmonary artery smooth muscle cells (RPASMCs) were cultured under normoxic conditions (5% CO2-95% ambient air) or hypoxic conditions (5% O2 for 6 h, 12 h, 24 h, 48 h). Hypoxic rats developed pulmonary arterial remodeling and right ventricular hypertrophy with significantly increased pulmonary arterial pressure. The levels of Nur77, HIF-1α and PNCA were upregulated in pulmonary arterial smooth muscle from hypoxic rats. Silencing of either Nur77 or HIF-1α attenuated hypoxia-induced proliferation. Silencing of HIF-1α down-regulated Nur77 protein level, but Nur77 silence did not reduce HIF-1α. Nur77 was not con-immunoprecipitated with HIF-1α. This study demonstrated that Nur77 acted as a downstream regulator of HIF-1α under hypoxia, and plays a critical role in the hypoxia-induced pulmonary vascular remodeling, which is regulated by HIF-1α. Nur77 maybe a novel target of HPH therapy.

Does Cell-Type-Specific Silencing of Monoamine Oxidase B Interfere with the Development of Right Ventricle (RV) Hypertrophy or Right Ventricle Failure in Pulmonary Hypertension?

Increased mitochondrial reactive oxygen species (ROS) formation is important for the development of right ventricular (RV) hypertrophy (RVH) and failure (RVF) during pulmonary hypertension (PH). ROS molecules are produced in different compartments within the cell, with mitochondria known to produce the strongest ROS signal. Among ROS-forming mitochondrial proteins, outer-mitochondrial-membrane-located monoamine oxidases (MAOs, type A or B) are capable of degrading neurotransmitters, thereby producing large amounts of ROS. In mice, MAO-B is the dominant isoform, which is present in almost all cell types within the heart. We analyzed the effect of an inducible cardiomyocyte-specific knockout of MAO-B (cmMAO-B KO) for the development of RVH and RVF in mice. Right ventricular hypertrophy was induced by pulmonary artery banding (PAB). RV dimensions and function were measured through echocardiography. ROS production (dihydroethidium staining), protein kinase activity (PamStation device), and systemic hemodynamics (in vivo catheterization) were assessed. A significant decrease in ROS formation was measured in cmMAO-B KO mice during PAB compared to Cre-negative littermates, which was associated with reduced activity of protein kinases involved in hypertrophic growth. In contrast to littermates in which the RV was dilated and hypertrophied following PAB, RV dimensions were unaffected in response to PAB in cmMAO-B KO mice, and no decline in RV systolic function otherwise seen in littermates during PAB was measured in cmMAO-B KO mice. In conclusion, cmMAO-B KO mice are protected against RV dilatation, hypertrophy, and dysfunction following RV pressure overload compared to littermates. These results support the hypothesis that cmMAO-B is a key player in causing RV hypertrophy and failure during PH.

Sex-dependent remodeling of right ventricular function in a rat model of pulmonary arterial hypertension.

Right ventricular (RV) function is an important prognostic indicator for pulmonary arterial hypertension (PAH), a vasculopathy that primarily and disproportionally affects women with distinct pre- and postmenopausal clinical outcomes. However, most animal studies have overlooked the impact of sex and ovarian hormones on RV remodeling in PAH. Here, we combined invasive measurements of RV hemodynamics and morphology with computational models of RV biomechanics in sugen-hypoxia (SuHx)-treated male, ovary-intact female, and ovariectomized female rats. Despite similar pressure overload levels, SuHx induced increases in end-diastolic elastance and passive myocardial stiffening, notably in male SuHx animals, corresponding to elevated diastolic intracellular calcium. Increases in end-systolic chamber elastance were largely explained by myocardial hypertrophy in male and ovary-intact female rats, whereas ovariectomized females exhibited contractility recruitment via calcium transient augmentation. Ovary-intact female rats primarily responded with hypertrophy, showing fewer myocardial mechanical alterations and less stiffening. These findings highlight sex-related RV remodeling differences in rats, affecting systolic and diastolic RV function in PAH.NEW & NOTEWORTHY Combining hemodynamic and morphological measurements from male, female, and ovariectomized female pulmonary arterial hypertension (PAH) rats revealed distinct adaptation mechanisms despite similar pressure overload. Males showed the most diastolic stiffening. Ovariectomized females had enhanced myocyte contractility and calcium transient upregulation. Ovary-intact females primarily responded with hypertrophy, experiencing milder passive myocardial stiffening and no changes in myocyte shortening. These findings suggest potential sex-specific pathways in right ventricular (RV) adaptation to PAH, with implications for targeted interventions.

Forsythoside B Mitigates Monocrotaline-Induced Pulmonary Arterial Hypertension via Blocking the NF-κB Signaling Pathway to Attenuate Vascular Remodeling.

Purpose: Pulmonary arterial hypertension (PAH) is a devastating disease with little effective treatment. The proliferation of pulmonary artery smooth muscle cells (PASMCs) induced by the nuclear factor-κB (NF-κB) signaling activation plays a pivotal role in the pathogenesis of PAH. Forsythoside B (FTS•B) possesses inhibitory effect on NF-κB signaling pathway. The present study aims to explore the effects and mechanisms of FTS•B in PAH. Methods: Sprague-Dawley rats received monocrotaline (MCT) intraperitoneal injection to establish PAH model, and FTS•B was co-treated after MCT injection. Right ventricular hypertrophy and pulmonary artery pressure were measured by echocardiography and right heart catheterization, respectively. Histological alterations were detected by H&E staining and immunohistochemistry. FTS•B's role in PASMC proliferation and migration were evaluated by CCK-8 and wound healing assay. To investigate the underlying mechanisms, Western blotting, immunofluorescence staining and ELISA were conducted. The NF-κB activator PMA was used to investigate the role of NF-κB in FTS•B's protective effects against PAH. Results: FTS•B markedly alleviated MCT-induced vascular remodeling and pulmonary artery pressure, and improved right ventricular hypertrophy and survival. FTS•B also reversed PDGF-BB-induced PASMC proliferation and migration, decreased PCNA and CyclinD1 expression in vitro. The elevated levels of IL-1ß and IL-6 caused by MCT were decreased by FTS•B. Mechanistically, MCT-triggered phosphorylation of p65, IκBα, IKKα and IKKß was blunted by FTS•B. FTS•B also reversed MCT-induced nuclear translocation of p65. However, all these protective effects were blocked by PMA-mediated NF-κB activation. Conclusion: FTS•B effectively attenuates PAH by suppressing the NF-κB signaling pathway to attenuate vascular remodeling. FTS•B might be a promising drug candidate with clinical translational potential for the treatment of PAH.

Multi-omic and multispecies analysis of right ventricular dysfunction.

BACKGROUND: Right ventricular failure (RVF) is a leading cause of morbidity and mortality in multiple cardiovascular diseases, but there are no treatments for RVF as therapeutic targets are not clearly defined. Contemporary transcriptomic/proteomic evaluations of RVF are predominately conducted in small animal studies, and data from large animal models are sparse. Moreover, a comparison of the molecular mediators of RVF across species is lacking. METHODS: Transcriptomics and proteomics analyses defined the pathways associated with cardiac magnetic resonance imaging (MRI)-derived values of RV hypertrophy, dilation, and dysfunction in control and pulmonary artery banded (PAB) pigs. Publicly available data from rat monocrotaline-induced RVF and pulmonary arterial hypertension patients with preserved or impaired RV function were used to compare molecular responses across species. RESULTS: PAB pigs displayed significant right ventricle/ventricular (RV) hypertrophy, dilation, and dysfunction as quantified by cardiac magnetic resonance imaging. Transcriptomic and proteomic analyses identified pathways associated with RV dysfunction and remodeling in PAB pigs. Surprisingly, disruptions in fatty acid oxidation (FAO) and electron transport chain (ETC) proteins were different across the 3 species. FAO and ETC proteins and transcripts were mostly downregulated in rats but were predominately upregulated in PAB pigs, which more closely matched the human response. All species exhibited similar dysregulation of the dilated cardiomyopathy and arrhythmogenic right ventricular cardiomyopathy pathways. CONCLUSIONS: The porcine metabolic molecular signature was more similar to human RVF than rodents. These data suggest there may be divergent molecular responses of RVF across species, and pigs may more accurately recapitulate metabolic aspects of human RVF.

Exploring the involvement of TASK-1 in the control of isolated rat right atrium function from healthy animals and an experimental model of monocrotaline-induced pulmonary hypertension.

The TASK-1 channel belongs to the two-pore domain potassium channel family. It is expressed in several cells of the heart, including the right atrial (RA) cardiomyocytes and the sinus node, and TASK-1 channel has been implicated in the pathogenesis of atrial arrhythmias (AA). Thus, using the rat model of monocrotaline-induced pulmonary hypertension (MCT-PH), we explored the involvement of TASK-1 in AA. Four-week-old male Wistar rats were injected with 50 mg/kg of MCT to induce MCT-PH and isolated RA function was studied 14 days later. Additionally, isolated RA from six-week-old male Wistar rats were used to explore the ability of ML365, a selective blocker of TASK-1, to modulate RA function. The hearts developed right atrial and ventricular hypertrophy, inflammatory infiltrate and the surface ECG demonstrated increased P wave duration and QT interval, which are markers of MCT-PH. The isolated RA from the MCT animals showed enhanced chronotropism, faster contraction and relaxation kinetics, and a higher sensibility to extracellular acidification. However, the addition of ML365 to extracellular media was not able to restore the phenotype. Using a burst pacing protocol, the RA from MCT animals were more susceptible to develop AA, and simultaneous administration of carbachol and ML365 enhanced AA, suggesting the involvement of TASK-1 in AA induced by MCT. TASK-1 does not play a key role in the chronotropism and inotropism of healthy and diseased RA; however, it may play a role in AA in the MCT-PH model.

Prostaglandin E1 reduces apoptosis and improves the homing of mesenchymal stem cells in pulmonary arterial hypertension by regulating hypoxia-inducible factor 1 alpha.

BACKGROUND: Pulmonary arterial hypertension (PAH) is associated with oxidative stress and affects the survival and homing of transplanted mesenchymal stem cells (MSCs) as well as cytokine secretion by the MSCs, thereby altering their therapeutic potential. In this study, we preconditioned the MSCs with prostaglandin E1 (PGE1) and performed in vitro and in vivo cell experiments to evaluate the therapeutic effects of MSCs in rats with PAH. METHODS: We studied the relationship between PGE1 and vascular endothelial growth factor (VEGF) secretion, B-cell lymphoma 2 (Bcl-2) expression, and C-X-C chemokine receptor 4 (CXCR4) expression in MSCs and MSC apoptosis as well as migration through the hypoxia-inducible factor (HIF) pathway in vitro. The experimental rats were randomly divided into five groups: (I) control group, (II) monocrotaline (MCT) group, (III) MCT + non-preconditioned (Non-PC) MSC group, (IV) MCT + PGE1-preconditioned (PGE1-PC) MSC group, and (V) MCT+PGE1+YC-1-PCMSC group. We studied methane dicarboxylic aldehyde (MDA) levels, MSC homing to rat lungs, mean pulmonary artery pressure, pulmonary artery systolic pressure, right ventricular hypertrophy index, wall thickness index (%WT), and relative wall area index (%WA) of rat pulmonary arterioles. RESULTS: Preconditioning with PGE1 increased the protein levels of HIF-1 alpha (HIF-1α) in MSCs, which can reduce MSC apoptosis and increase the protein levels of CXCR4, MSC migration, and vascular endothelial growth factor secretion. Upon injection with PGE1-PCMSCs, the pulmonary artery systolic pressure, mean pulmonary artery pressure, right ventricular hypertrophy index, %WT, and %WA decreased in rats with PAH. PGE1-PCMSCs exhibited better therapeutic effects than non-PCMSCs. Interestingly, lificiguat (YC-1), an inhibitor of the HIF pathway, blocked the effects of PGE1 preconditioning. CONCLUSIONS: Our findings indicate that PGE1 modulates the properties of MSCs by regulating the HIF pathway, providing insights into the mechanism by which PGE1 preconditioning can be used to improve the therapeutic potential of MSCs in PAH.

Bioactive Compounds From Coptidis Rhizoma Alleviate Pulmonary Arterial Hypertension by Inhibiting Pulmonary Artery Smooth Muscle Cells' Proliferation and Migration.

ABSTRACT: Pulmonary arterial hypertension (PAH) is a devastating disorder characterized by excessive proliferation and vasoconstriction of small pulmonary artery vascular smooth muscle cells (PASMCs). Coptidis rhizoma (CR) because of the complexity of the components, the underlying pharmacological role and mechanism of it on PAH remains unknown. In this article, the network pharmacological analysis was used to screen the main active constituents of CR and the molecular targets that these constituents act on. Then, we evaluated the importance of berberine and quercetin (biologically active components of CR) on the proliferation and migration of PASMCs and vascular remodeling in experimental models of PAH. Our results showed that berberine and quercetin effectively inhibited the proliferation and migration of hypoxia-induced PASMCs in a manner likely to be mediated by the suppression of MAPK1, NADPH oxidase 4 (NOX4), and cytochrome P450 1B1 (CYP1B1) expression. Furthermore, berberine and quercetin treatment attenuates pulmonary hypertension, reduces right ventricular hypertrophy, and improves pulmonary artery remodeling in monocrotaline-induced pulmonary hypertension in rat models. In conclusion, this research demonstrates CR might be a promising treatment option for PAH, and the network pharmacology approach can be an effective tool to reveal the potential mechanisms of Chinese herbal medicine.

Thioredoxin system activation is associated with the progression of experimental pulmonary arterial hypertension.

In addition to being an antioxidant, thioredoxin (Trx) is known to stimulate signaling pathways involved in cell proliferation and to inhibit apoptosis. The aim of this study was to explore the role of Trx in some of these pathways along the progression of monocrotaline (MCT)-induced pulmonary arterial hypertension (PAH). Male rats were first divided into two groups: monocrotaline (MCT - 60 mg/kg i.p.) and control (received saline), that were further divided into three groups: 1, 2, and 3 weeks. Animals were submitted to echocardiographic analysis. Right and left ventricles were used for the measurement of hypertrophy, through morphometric and histological analysis. The lung was prepared for biochemical and molecular analysis. One week after MCT injection, there was an increase in thioredoxin reductase (TrxR) activity, a reduction in glutathione reductase (GR) activity, and an increase in Trx-1 and vitamin D3 up-regulated protein-1 (VDUP-1) expression. Two weeks after MCT injection, there was an increase in VDUP-1, Akt and cleaved caspase-3 activation, and a decrease in Trx-1 and Nrf2 expression. PAH-induced by MCT promoted a reduction in Nrf2 and Trx-1 expression as well as an increase in Akt and VDUP-1 expression after three weeks. The increase in pulmonary vascular resistance was accompanied by increased TrxR activity, suggesting an association between the Trx system and functional changes in the progression of PAH. It seems that Trx-1 activation was an adaptive response to MCT administration to cope with pulmonary remodeling and disease progression, suggesting a potential new target for PAH therapeutics.

Right-sided heart failure is also associated with transverse tubule remodeling in the left ventricle.

Right-sided heart failure is a common consequence of pulmonary arterial hypertension. Overloading the right ventricle results in right ventricular hypertrophy, which progresses to failure in a process characterized by impaired Ca2+ dynamics and force production that is linked with transverse (t)-tubule remodeling. This also unloads the left ventricle, which consequently atrophies. Experimental left-ventricular unloading can result in t-tubule remodeling, but it is currently unclear if this occurs in right-sided heart failure. In this work, we used a model of monocrotaline (MCT)-induced right heart failure in male rats, using confocal microscopy to investigate cellular remodeling of t-tubules, junctophilin-2 (JPH2), and ryanodine receptor-2 (RyR2). We examined remodeling across tissue anatomical regions of both ventricles: in trabeculae, papillary muscles, and free walls. Our analyses revealed that MCT hearts demonstrated a significant loss of t-tubule periodicity, disruption of the normal sarcomere striated pattern with JPH2 labeling, and also a disorganized striated pattern of RyR2, a feature not previously reported in right heart failure. Remodeling of JPH2 and RyR2 in the MCT heart was more pronounced in papillary muscles and trabeculae compared with free walls, particularly in the left ventricle. We find that these structures, commonly used as ex vivo muscle preparations, are more sensitive to the disease process.NEW & NOTEWORTHY In this work, we demonstrate that t-tubule remodeling occurs in the atrophied left ventricle as well as the overloaded right ventricle after right-side heart failure. Moreover, we identify that t-tubule remodeling in both ventricles is linked to sarcoplasmic reticulum remodeling as indicated by decreased labeling periodicity of both the Ca2+ release channel, RyR2, and the cardiac junction-forming protein, JPH2, that forms a link between the sarcoplasmic reticulum and sarcolemma. Studies developing treatments for right-sided heart failure should consider effects on both the right and left ventricle.

Distinct time courses and mechanics of right ventricular hypertrophy and diastolic stiffening in a male rat model of pulmonary arterial hypertension.

Although pulmonary arterial hypertension (PAH) leads to right ventricle (RV) hypertrophy and structural remodeling, the relative contributions of changes in myocardial geometric and mechanical properties to systolic and diastolic chamber dysfunction and their time courses remain unknown. Using measurements of RV hemodynamic and morphological changes over 10 wk in a male rat model of PAH and a mathematical model of RV mechanics, we discriminated the contributions of RV geometric remodeling and alterations of myocardial material properties to changes in systolic and diastolic chamber function. Significant and rapid RV hypertrophic wall thickening was sufficient to stabilize ejection fraction in response to increased pulmonary arterial pressure by week 4 without significant changes in systolic myofilament activation. After week 4, RV end-diastolic pressure increased significantly with no corresponding changes in end-diastolic volume. Significant RV diastolic chamber stiffening by week 5 was not explained by RV hypertrophy. Instead, model analysis showed that the increases in RV end-diastolic chamber stiffness were entirely attributable to increased resting myocardial material stiffness that was not associated with significant myocardial fibrosis or changes in myocardial collagen content or type. These findings suggest that whereas systolic volume in this model of RV pressure overload is stabilized by early RV hypertrophy, diastolic dilation is prevented by subsequent resting myocardial stiffening.NEW & NOTEWORTHY Using a novel combination of hemodynamic and morphological measurements over 10 wk in a male rat model of PAH and a mathematical model of RV mechanics, we found that compensated systolic function was almost entirely explained by RV hypertrophy, but subsequently altered RV end-diastolic mechanics were primarily explained by passive myocardial stiffening that was not associated with significant collagen extracellular matrix accumulation.

Interleukin-17 aggravates right ventricular remodeling via activating STAT3 under both normoxia and hypoxia.

OBJECTIVE: Proinflammatory cytokine interleukin 17 (IL-17) is involved in ventricular remodeling, mainly of the left ventricle. This study was designed to explore the role of IL-17 played in the pathogenesis of right ventricular hypertrophy (RVH), aiming to provide a novel treatment target or diagnostic biomarker options for improving the care of RVH patients. METHODS: C57BL/6 mice were maintained in 10% O2 chamber or room air for four weeks. Right ventricular hypertrophy index (RVHI), RV/body weight ratio, pulmonary arteriolar remodeling determined by percent media thickness (%MT), and the cardiomyocyte diameter of RV were evaluated. Mice were treated with exogenous recombinant mouse IL-17 (rmIL-17, 1 µg per dose twice a week) for four weeks. H9c2 cardiomyocytes were cultured and treated with IL-17 (10 ng/mL) and STAT3 inhibitor (10 ng/mL) either under normoxia (21% O2, 5% CO2, 74% N2) or under hypoxia (3% O2, 5% CO2, 92% N2). Cardiomyocyte viability was assessed by Cell counting kit 8 (CCK-8) assay. The mRNA level was detected by RT-PCR, where as the protein expression was measured by Western blot, immunohistochemistry, and immunofluorescent analyses. RESULTS: In vivo experiments showed that IL-17 did not affect the pulmonary artery under normoxia, after treatment with rmIL-17, %MT was not changed, while RVHI and the RV/body weight ratio were increased, indicating that IL-17 directly induced right ventricular hypertrophy. In a time-course study, the mice were exposed to hypoxia for 0, 1, 2, 3, 4 weeks, respectively. We found that the expression of IL-17 was gradually upregulated in RV tissue in a time-dependent manner after one week of hypoxia exposure, especially at the third and fourth week. Cardiomyocyte hypertrophy and apoptosis were observed after the exposure of the mice to hypoxia for four weeks, rmIL-17 further aggravated the hypoxia-induced cardiomyocyte hypertrophy and apoptosis. The expression of p-STAT3 in the IL-17-deficient mice was lower than in the wild-type mice. In vitro, IL-17 inhibited cardiomyocyte viability and induced cardiomyocyte apoptosis via STAT3 under both normoxic and hypoxic conditions. CONCLUSIONS: These findings support a role for IL-17 as a mediator in the pathogenesis RVH, which might be considered as a potential novel anti-inflammation therapeutic strategy or diagnostic biomarker for RVH.

CENPE contributes to pulmonary vascular remodeling in pulmonary hypertension.

Hypoxic pulmonary vascular remodeling is a pathological feature of pulmonary hypertension (PH). Our results showed that centromere-associated protein E (CENPE) expression in PH patients and hypoxia-induced PH rats was significantly higher than that in normal controls. In addition, CENPE deficiency significantly inhibited the development of pulmonary vascular remodeling and right ventricular hypertrophy. Moreover, knocking out CENPE effectively inhibited the proliferation and induced the apoptosis of primary pulmonary artery smooth muscle cells (PASMCs) in vivo. Furthermore, CENPE silencing by small interference significantly inhibited abnormal proliferation, apoptosis resistance, migration, and cell cycle arrest in hypoxia-induced PASMCs. Interestingly, we found that CENPE might exert its biological effect by targeting the transcription of CDK1 proteins.

Newer insights into the pathobiological and pharmacological basis of the sex disparity in patients with pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) affects more women than men, although affected females tend to survive longer than affected males. This sex disparity in PAH is postulated to stem from the diverse roles of sex hormones in disease etiology. In animal models, estrogens appear to be implicated not only in pathologic remodeling of pulmonary arteries, but also in protection against right ventricular (RV) hypertrophy. In contrast, the male sex hormone testosterone is associated with reduced survival in male animals, where it is associated with increased RV mass, volume, and fibrosis. However, it also has a vasodilatory effect on pulmonary arteries. Furthermore, patients of both sexes show varying degrees of response to current therapies for PAH. As such, there are many gaps and contradictions regarding PAH development, progression, and therapeutic interventions in male versus female patients. Many of these questions remain unanswered, which may be due in part to lack of effective experimental models that can consistently reproduce PAH pulmonary microenvironments in their sex-specific forms. This review article summarizes the roles of estrogens and related sex hormones, immunological and genetical differences, and the benefits and limitations of existing experimental tools to fill in gaps in our understanding of the sex-based variation in PAH development and progression. Finally, we highlight the potential of a new tissue chip-based model mimicking PAH-afflicted male and female pulmonary arteries to study the sex-based differences in PAH and to develop personalized therapies based on patient sex and responsiveness to existing and new drugs.

Up-regulation of circRNA_0068481 promotes right ventricular hypertrophy in PAH patients via regulating miR-646/miR-570/miR-885.

CircRNA-0068481 and several miRNAs are important in the pathogenesis of right ventricular hypertrophy (VH), while the inhibition of eye absent transcriptional coactivator and phosphatase 3 (EYA3) was proved to reverse vascular remodelling in rats. In this study, we tried to study the diagnostic value and mechanistic role of circRNA_0068481 in the diagnosis of RVH in PAH patients. qPCR was done to measure circRNA-0068481, miR-646, miR-750, miR-885 and EYA3 mRNA expression. Luciferase assay was done to explore the regulatory relationship between circRNA-0068481/EYA3 and the miRNAs. Western blot was done to measure EYA3 expression in AC16 cells. The expression of circRNA-0068481, miR-646 and miR-570 showed a considerable capability to diagnose RVH in PAH patients. The luciferase activity of circRNA-0068481 was remarkably suppressed by miR-646, miR-570 or miR-885. The luciferase signal of EYA3 was also inhibited by miR-646, miR-570 and miR-885. Up-regulation of circRNA-0068481 expression in AC16 significantly decreased miR-646, miR-570 and miR-885 expression, and up-regulated EYA3 expression, whereas circRNA-0068481 down-regulation significantly increased miR-646, miR-570 and miR-885 expression, and repressed EYA3 expression. CircRNA_0068481 sponged several miRNAs including miR-646, miR-570 and miR-885. These miRNAs were all found to target the expression of EYA3 mRNA, which is involved in the onset of right ventricular hypertrophy. Therefore, it can be concluded that the up-regulation of circRNA_0068481 can predict the diagnosis of right ventricular hypertrophy in pulmonary arterial hypertension patients.

Exercise blood pressure and cardiac structure: A systematic review and meta-analysis of cross-sectional studies.

OBJECTIVES: A hypertensive response to exercise (HRE) is associated with cardiovascular disease and high blood pressure (BP). Sub-clinical changes to cardiac structure may underlie these associations, although this has not been systematically determined. Via systematic review and meta-analysis, we aimed to (1) assess the relationship between exercise BP and cardiac structure, and (2) determine if cardiac structure is altered in those with an HRE, across various study populations (including those with/without high BP at rest). DESIGN AND METHODS: Three online databases were searched for cross-sectional studies reporting exercise BP, HRE and cardiac structural variables. Random-effects meta-analyses and meta-regressions were used to calculate pooled correlations between exercise BP and cardiac structure, and pooled mean differences and relative risk between those with/without an HRE. RESULTS: Forty-nine studies, (n=23,707 total; aged 44±4 years; 63% male) were included. Exercise systolic BP was associated with increased left ventricular (LV) mass, LV mass index, relative wall thickness, posterior wall thickness and interventricular septal thickness (p<0.05 all). Those with an HRE had higher risk of LV hypertrophy (relative risk: 2.6 [1.85-3.70]), increased LV mass (47±7g), LV mass index (7±2g/m2), relative wall thickness (0.02±0.005), posterior wall thickness (0.78±0.20mm), interventricular septal thickness (0.78±0.17mm) and left atrial diameter (2±0.52mm) vs. those without an HRE (p<0.05 all). Results were broadly similar between studies with different population characteristics. CONCLUSIONS: Exercise systolic BP is associated with cardiac structure, and those with an HRE show evidence towards adverse remodelling. Results were similar across different study populations, highlighting the hypertension-related cardiovascular risk associated with an HRE.

Angiotensin II type 1 receptor mediates pulmonary hypertension and right ventricular remodeling induced by inhaled nicotine.

Use of electronic cigarettes is rapidly increasing among youth and young adults, but little is known regarding the long-term cardiopulmonary health impacts of these nicotine-containing devices. Our group has previously demonstrated that chronic, inhaled nicotine induces pulmonary hypertension (PH) and right ventricular (RV) remodeling in mice. These changes were associated with upregulated RV angiotensin-converting enzyme (ACE). Angiotensin II receptor blockers (ARBs) have been shown to reverse cigarette smoking-induced PH in rats. ACE inhibitor and ARB use in a large retrospective cohort of patients with PH is associated with improved survival. Here, we utilized losartan (an ARB specific for angiotensin II type 1 receptor) to further explore nicotine-induced PH. Male C57BL/6 mice received nicotine vapor for 12 h/day, and exposure was assessed using serum cotinine to achieve levels comparable to human smokers or electronic cigarette users. Mice were exposed to nicotine for 8 wk and a subset was treated with losartan via an osmotic minipump. Cardiac function was assessed using echocardiography and catheterization. Although nicotine exposure increased angiotensin II in the RV and lung, this finding was nonsignificant. Chronic, inhaled nicotine significantly increased RV systolic pressure and RV free wall thickness versus air control. These parameters were significantly lower in mice receiving both nicotine and losartan. Nicotine significantly increased RV internal diameter, with no differences seen between the nicotine and nicotine-losartan group. Neither nicotine nor losartan affected left ventricular structure or function. These findings provide the first evidence that antagonism of the angiotensin II type 1 receptor can ameliorate chronic, inhaled nicotine-induced PH and RV remodeling.NEW & NOTEWORTHY Chronic, inhaled nicotine causes pulmonary hypertension and right ventricular remodeling in mice. Treatment with losartan, an angiotensin II type 1 receptor antagonist, ameliorates nicotine-induced pulmonary hypertension and right ventricular remodeling. This novel finding provides preclinical evidence for the use of renin-angiotensin system-based therapies in the treatment of pulmonary hypertension, particularly in patients with a history of tobacco-product use.