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The integration of data from various healthcare centers into disease registries is pivotal for facilitating collaborative research and enhancing clinical insights. In this study, we investigate the integration process of existing registries into the PVRI GoDeep meta-registry, focusing on the complexities and challenges encountered. We detail the integration process, including data transformation, mapping updates, and feedback mechanisms. Our findings underscore the importance of standardized processes and proactive communication in addressing data quality issues, ultimately enhancing the reliability and trustworthiness of meta-registry data. Through careful harmonization of the data and transparent documentation of data processing, we pave the way for leveraging registry data to drive advancements in pulmonary hypertension research and patient care.

Aims: Hepatic T1-time derived from cardiac magnetic resonance imaging (cMRI) reflects venous congestion and may provide a simple alternative to invasive end-diastolic elastance (Eed) for assessment of right ventricular (RV) diastolic function. We investigated the association of native hepatic T1-time with single-beat Eed and the value of hepatic T1-time for longitudinal monitoring in pulmonary hypertension (PH)., Methods and Results: We retrospectively enrolled 85 patients with suspected PH (59% female; 78 with PH diagnosed; 7 with PH excluded) who underwent standard right heart catheterization and cMRI within 24 h between 2015 and 2020. Hepatic T1-time showed moderate to strong correlations (rho >0.3, P ≤ 0.002) with pulmonary vascular resistance, native myocardial T1-time, Eed, RV size and function, brain natriuretic peptide (BNP) level, and 6-min walk distance, and a significant association with functional class (Kruskal-Wallis P < 0.001). Eed, myocardial T1-time, and BNP were independently linked to hepatic T1-time in multivariable regression. Hepatic T1-time > 598 ms predicted elevated Eed with 72.9% sensitivity and 82.1% specificity. Hepatic T1-time was superior to Eed in predicting clinical worsening. In 16 patients with follow-up assessments, those with decreasing hepatic T1-time (7 patients) showed significant hemodynamic improvements, whereas those with increasing hepatic T1-time (9 patients) did not. In a second retrospective cohort of 27 patients with chronic thromboembolic PH undergoing balloon pulmonary angioplasty, hepatic T1-time decreased significantly and hemodynamics improved after the procedure., Conclusions: Hepatic T1-time predicts RV diastolic dysfunction and prognosis, and may be useful for monitoring disease progression and treatment response in PH., Competing Interests: Declaration of competing interest NK has received speaking fees from Janssen outside the submitted work. AY has received personal fees from MSD outside the submitted work. WS has received speaker/consultancy fees from Pfizer and Bayer Pharma AG. CBW has received speaker fees and/or consultant honoraria from Actelion, AOP Orphan Pharmaceuticals AG, Bayer AG, BTG, MSD, and Pfizer. KT has received speaking fees from Actelion, Janssen, and Bayer. All other authors report no relationships that could be construed as a conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Pulmonary hypertension (PH) can lead to congestive hepatopathy, known as cardiohepatic syndrome (CHS). Hepatic congestion is associated with increased liver stiffness, which can be quantified using shear wave elastography. We aimed to investigate whether hepatic shear wave elastography detects patients at risk in the early stages of PH., Methods: Sixty-three prospectively enrolled patients undergoing right heart catheterization (52 diagnosed with PH and 11 with invasive exclusion of PH) and 52 healthy volunteers underwent assessments including echocardiography and hepatic shear wave elastography. CHS was defined as increased levels of ≥2 of the following: gamma-glutamyl transferase, alkaline phosphatase, and bilirubin. Liver stiffness was defined as normal (≤5.0 kPa) or high (>5.0 kPa)., Results: Compared with normal liver stiffness, high liver stiffness was associated with impaired right ventricular (RV) and right atrial (RA) function (median [interquartile range] RV ejection fraction: 54 [49; 57]% vs 45 [34; 51]%, p < 0.001; RA reservoir strain: 49 [41; 54]% vs 33 [22; 41]%, p < 0.001), more severe tricuspid insufficiency (p < 0.001), and higher prevalence of hepatovenous backflow (2% vs 29%, p < 0.001) and CHS (2% vs 10%, p = 0.038). In the patient subgroup with precapillary PH (n = 48), CHS and high liver stiffness were associated with increased European Society of Cardiology/European Respiratory Society 2022 risk scores (p = 0.003)., Conclusions: Shear wave liver elastography yields important information regarding right heart function and may complement risk assessment in patients with (suspected) PH., (Copyright © 2024 International Society for the Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Impaired respiratory variation of right atrial pressure (RAP) in severe pulmonary hypertension (PH) suggests difficulty tolerating increased preload during inspiration. Our study explores whether this impairment links to specific factors: right ventricular (RV) diastolic function, elevated RV afterload, systolic RV function, or RV-pulmonary arterial (PA) coupling. We retrospectively evaluated respiratory RAP variation in all participants enrolled in the EXERTION study. Impaired respiratory variation was defined as end-expiratory RAP - end-inspiratory RAP ≤ 2 mm Hg. RV function and afterload were evaluated using conductance catheterization. Impaired diastolic RV function was defined as end-diastolic elastance (Eed) ≥ median (0.19 mm Hg/mL). Seventy-five patients were included; PH was diagnosed in 57 patients and invasively excluded in 18 patients. Of the 75 patients, 31 (41%) had impaired RAP variation, which was linked with impaired RV systolic function and RV-PA coupling and increased tricuspid regurgitation and Eed as compared to patients with preserved RAP variation. In backward regression, RAP variation associated only with Eed. RAP variation but not simple RAP identified impaired diastolic RV function (area under the receiver operating characteristic curve [95% confidence interval]: 0.712 [0.592, 0.832] and 0.496 [0.358, 0.634], respectively). During exercise, patients with impaired RAP variation experienced greater RV dilatation and reduced diastolic reserve and cardiac output/index compared with patients with preserved RAP variation. Preserved RAP variation was associated with a better prognosis than impaired RAP variation based on the 2022 European Society of Cardiology/European Respiratory Society risk score (chi-square P = 0.025) and survival free from clinical worsening (91% vs 71% at 1 year and 79% vs 50% at 2 years [log-rank P = 0.020]; hazard ratio: 0.397 [95% confidence interval: 0.178, 0.884]). Subgroup analyses in patients with group 1 and group 4 PH demonstrated consistent findings with those observed in the overall study cohort. Respiratory RAP variations reflect RV diastolic function, are independent of RV-PA coupling or tricuspid regurgitation, are associated with exercise-induced haemodynamic changes, and are prognostic in PH.Trial registration. NCT04663217., (© 2024. The Author(s).)

Background: The precise origin of newly formed ACTA2+ (alpha smooth muscle actin-positive) cells appearing in nonmuscularized vessels in the context of pulmonary hypertension is still debatable although it is believed that they predominantly derive from preexisting vascular smooth muscle cells (VSMCs)., Methods: Gli1 Cre-ERT2 ; tdTomato flox mice were used to lineage trace GLI1+ (glioma-associated oncogene homolog 1-positive) cells in the context of pulmonary hypertension using 2 independent models of vascular remodeling and reverse remodeling: hypoxia and cigarette smoke exposure. Hemodynamic measurements, right ventricular hypertrophy assessment, flow cytometry, and histological analysis of thick lung sections followed by state-of-the-art 3-dimensional reconstruction and quantification using Imaris software were used to investigate the contribution of GLI1+ cells to neomuscularization of the pulmonary vasculature., Results: The data show that GLI1+ cells are abundant around distal, nonmuscularized vessels during steady state, and this lineage contributes to around 50% of newly formed ACTA2+ cells around these normally nonmuscularized vessels. During reverse remodeling, cells derived from the GLI1+ lineage are largely cleared in parallel to the reversal of muscularization. Partial ablation of GLI1+ cells greatly prevented vascular remodeling in response to hypoxia and attenuated the increase in right ventricular systolic pressure and right heart hypertrophy. Single-cell RNA sequencing on sorted lineage-labeled GLI1+ cells revealed an Acta2 high fraction of cells with pathways in cancer and MAPK (mitogen-activated protein kinase) signaling as potential players in reprogramming these cells during vascular remodeling. Analysis of human lung-derived material suggests that GLI1 signaling is overactivated in both group 1 and group 3 pulmonary hypertension and can promote proliferation and myogenic differentiation., Conclusions: Our data highlight GLI1+ cells as an alternative cellular source of VSMCs in pulmonary hypertension and suggest that these cells and the associated signaling pathways represent an important therapeutic target for further studies., Competing Interests: Disclosures None.

Background: COPD is an incurable disease and a leading cause of death worldwide. In mice, fibroblast growth factor (FGF)10 is essential for lung morphogenesis, and in humans, polymorphisms in the human FGF10 gene correlate with an increased susceptibility to develop COPD., Methods: We analysed FGF10 signalling in human lung sections and isolated cells from healthy donor, smoker and COPD lungs. The development of emphysema and PH was investigated in Fgf10 +/- and Fgfr2b +/- (FGF receptor 2b) mice upon chronic exposure to cigarette smoke. In addition, we overexpressed FGF10 in mice following elastase- or cigarette smoke-induced emphysema and pulmonary hypertension (PH)., Results: We found impaired FGF10 expression in human lung alveolar walls and in primary interstitial COPD lung fibroblasts. In contrast, FGF10 expression was increased in large pulmonary vessels in COPD lungs. Consequently, we identified impaired FGF10 signalling in alveolar walls as an integral part of the pathomechanism that leads to emphysema and PH development: mice with impaired FGF10 signalling ( Fgf10 +/- and Fgfr2b +/- ) spontaneously developed lung emphysema, PH and other typical pathomechanistic features that generally arise in response to cigarette smoke exposure., Conclusion: In a therapeutic approach, FGF10 overexpression successfully restored lung alveolar and vascular structure in mice with established cigarette smoke- and elastase-induced emphysema and PH. FGF10 treatment triggered an initial increase in the number of alveolar type 2 cells that gradually returned to the basal level when the FGF10-mediated repair process progressed. Therefore, the application of recombinant FGF10 or stimulation of the downstream signalling cascade might represent a novel therapeutic strategy in the future., Competing Interests: Conflict of interest: The authors have no potential conflicts of interest to disclose., (Copyright ©The authors 2023.)

Background: Fibroblast growth factor 23 (FGF-23) has been associated with left ventricular hypertrophy (LVH) and heart failure. However, its role in right ventricular (RV) remodeling and RV failure is unknown. This study analyzed the utility of FGF-23 as a biomarker of RV function in patients with pulmonary hypertension (PH)., Methods: In this observational study, FGF-23 was measured in the plasma of patients with PH (n = 627), dilated cardiomyopathy (DCM, n = 59), or LVH with severe aortic stenosis (n = 35). Participants without LV or RV abnormalities served as controls (n = 36)., Results: Median FGF-23 plasma levels were higher in PH patients than in healthy controls (p < 0.001). There were no significant differences between PH, DCM, and LVH patients. Analysis across tertiles of FGF-23 levels in PH patients revealed an association between higher FGF-23 levels and higher levels of NT-proBNP and worse renal function. Furthermore, patients in the high-FGF-23 tertile had a higher pulmonary vascular resistance (PVR), mean pulmonary artery pressure, and right atrial pressure and a lower cardiac index (CI) than patients in the low tertile (p < 0.001 for all comparisons). Higher FGF-23 levels were associated with higher RV end-diastolic diameter and lower tricuspid annular plane systolic excursions (TAPSE) and TAPSE/PASP. Receiver operating characteristic analysis revealed FGF-23 as a good predictor of RV maladaptation, defined as TAPSE < 17 mm and CI < 2.5 L/min/m 2 . Association of FGF-23 with parameters of RV function was independent of the glomerular filtration rate in regression analysis., Conclusion: FGF-23 may serve as a biomarker for maladaptive RV remodeling in patients with PH., (© 2023. The Author(s).)

Background: Right atrial (RA) imaging has emerged as a promising tool for the evaluation of patients with pulmonary hypertension (PH), albeit without systematic validation., Methods: PubMed, Web of Science and the Cochrane library were searched for studies investigating the prognostic value of RA imaging assessment in patients with PH from 2000 to June 2021 (PROSPERO Identifier: CRD42020212850). An inverse variance-weighted meta-analysis of univariable hazard ratios (HRs) was performed using a random effects model., Results: Thirty-five studies were included (3,476 patients with PH; 74% female, 86% pulmonary arterial hypertension). Risk of bias was low/moderate (Quality of Prognosis Studies checklist). RA area (HR 1.06; 95% confidence interval [CI] 1.04-1.08), RA indexed area (HR 1.09; 95% CI 1.04-1.14), RA peak longitudinal strain (PLS; HR 0.94; 95% CI 0.91-0.97) and RA total emptying fraction (HR 0.96; 95% CI 0.94-0.98) were significantly associated with combined end-points including death, clinical worsening and/or lung transplantation; RA volume and volume index showed marginal significant associations. RA area (HR 1.06; 95% CI 1.04-1.07), RA indexed area (HR 1.12; 95% CI 1.07-1.17) and RA PLS (HR 0.98; 95% CI 0.97-0.99) showed significant associations with mortality; RA total emptying fraction showed a marginal association., Conclusions: Imaging-based RA assessment qualifies as a relevant prognostic marker in PH. RA area reliably predicts composite end-points and mortality, which underscores its clinical utility. RA PLS emerged as a promising imaging measure, but is currently limited by the number of studies and different acquisition methods., (Copyright © 2022 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Background: This study analyzed the utility of soluble ST2 (sST2) and GDF-15 as biomarkers of right ventricular (RV) function in patients with pulmonary hypertension (PH). Methods: GDF-15 and sST2 serum concentrations were measured in patients with PH (n = 628), dilated cardiomyopathy (n = 31) and left ventricular hypertrophy (n = 47), and in healthy controls (n = 61). Results: Median sST2 and GDF-15 levels in patients with left ventricular hypertrophy were higher than in patients with PH and dilated cardiomyopathy. In tertile analysis GDF-15 >1363 pg/ml and sST2 >38 ng/ml were associated with higher N-terminal pro-brain natriuretic peptide, RV systolic dysfunction, RV-pulmonary arterial uncoupling and hemodynamic impairment. Conclusion: GDF-15 and sST2 are potential biomarkers of RV dysfunction in patients with PH.

Background: Pulmonary arterial hypertension (PAH) is a progressive disease characterised by pro-proliferative and anti-apoptotic phenotype in vascular cells, leading to pulmonary vascular remodelling and right heart failure. Peptidyl-prolyl cis / trans isomerase, NIMA interacting 1 (Pin1), a highly conserved enzyme, which binds to and catalyses the isomerisation of specific phosphorylated Ser/Thr-Pro motifs, acts as a molecular switch in multiple coordinated cellular processes. We hypothesised that Pin1 plays a substantial role in PAH, and its inhibition with a natural organic compound, Juglone, would reverse experimental pulmonary hypertension., Results: We demonstrated that the expression of Pin1 was markedly elevated in experimental pulmonary hypertension ( i.e. hypoxia-induced mouse and Sugen/hypoxia-induced rat models) and pulmonary arterial smooth muscle cells of patients with clinical PAH. In vitro Pin1 inhibition by either Juglone treatment or short interfering RNA knockdown resulted in an induction of apoptosis and decrease in proliferation of human pulmonary vascular cells. Stimulation with growth factors induced Pin1 expression, while its inhibition reduced the activity of numerous PAH-related transcription factors, such as hypoxia-inducible factor (HIF)-α and signal transducer and activator of transcription (STAT). Juglone administration lowered pulmonary vascular resistance, enhanced right ventribular function, improved pulmonary vascular and cardiac remodelling in the Sugen/hypoxia rat model of PAH and the chronic hypoxia-induced pulmonary hypertension model in mice., Conclusion: Our study demonstrates that targeting of Pin1 with small molecule inhibitor, Juglone, might be an attractive future therapeutic strategy for PAH and right heart disease secondary to PAH., Competing Interests: Conflict of interest: W. Seeger reports personal fees from Actelion, Bayer AG, Novartis, Vectura, Medspray and United Therapeutics, outside the submitted work. All other authors disclose no potential conflicts of interest., (Copyright ©The authors 2022.)

Obesity is a pre-disposing condition for chronic obstructive pulmonary disease, asthma, and pulmonary arterial hypertension. Accumulating evidence suggests that metabolic influences during development can determine chronic lung diseases (CLD). We demonstrate that maternal obesity causes early metabolic disorder in the offspring. Here, interleukin-6 induced bronchial and microvascular smooth muscle cell (SMC) hyperproliferation and increased airway and pulmonary vascular resistance. The key anti-proliferative transcription factor FoxO1 was inactivated via nuclear exclusion. These findings were confirmed using primary SMC treated with interleukin-6 and pharmacological FoxO1 inhibition as well as genetic FoxO1 ablation and constitutive activation. In vivo, we reproduced the structural and functional alterations in offspring of obese dams via the SMC-specific ablation of FoxO1. The reconstitution of FoxO1 using IL-6-deficient mice and pharmacological treatment did not protect against metabolic disorder but prevented SMC hyperproliferation. In human observational studies, childhood obesity was associated with reduced forced expiratory volume in 1 s/forced vital capacity ratio Z-score (used as proxy for lung function) and asthma. We conclude that the interleukin-6-FoxO1 pathway in SMC is a molecular mechanism by which perinatal obesity programs the bronchial and vascular structure and function, thereby driving CLD development. Thus, FoxO1 reconstitution provides a potential therapeutic option for preventing this metabolic programming of CLD., (© 2022. The Author(s).)

Phenotypic alterations in resident vascular cells contribute to the vascular remodeling process in diseases such as pulmonary (arterial) hypertension [P(A)H]. How the molecular interplay between transcriptional coactivators, transcription factors (TFs), and chromatin state alterations facilitate the maintenance of persistently activated cellular phenotypes that consequently aggravate vascular remodeling processes in PAH remains poorly explored. RNA sequencing (RNA-seq) in pulmonary artery fibroblasts (FBs) from adult human PAH and control lungs revealed 2460 differentially transcribed genes. Chromatin immunoprecipitation sequencing (ChIP-seq) revealed extensive differential distribution of transcriptionally accessible chromatin signatures, with 4152 active enhancers altered in PAH-FBs. Integrative analysis of RNA-seq and ChIP-seq data revealed that the transcriptional signatures for lung morphogenesis were epigenetically derepressed in PAH-FBs, including coexpression of T-box TF 4 ( TBX4 ), TBX5 , and SRY-box TF 9 ( SOX9 ), which are involved in the early stages of lung development. These TFs were expressed in mouse fetuses and then repressed postnatally but were maintained in persistent PH of the newborn and reexpressed in adult PAH. Silencing of TBX4 , TBX5 , SOX9 , or E1A-associated protein P300 ( EP300 ) by RNA interference or small-molecule compounds regressed PAH phenotypes and mesenchymal signatures in arterial FBs and smooth muscle cells. Pharmacological inhibition of the P300/CREB-binding protein complex reduced the remodeling of distal pulmonary vessels, improved hemodynamics, and reversed established PAH in three rodent models in vivo, as well as reduced vascular remodeling in precision-cut tissue slices from human PAH lungs ex vivo. Epigenetic reactivation of TFs associated with lung development therefore underlies PAH pathogenesis, offering therapeutic opportunities.

Background: Pulmonary hypertension (PH) is a common complication of COPD, associated with increased mortality and morbidity. Intriguingly, pulmonary vascular alterations have been suggested to drive emphysema development. Previously, we identified inducible nitric oxide synthase (iNOS) as an essential enzyme for development and reversal of smoke-induced PH and emphysema, and showed that iNOS expression in bone-marrow-derived cells drives pulmonary vascular remodelling, but not parenchymal destruction. In this study, we aimed to identify the iNOS-expressing cell type driving smoke-induced PH and to decipher pro-proliferative pathways involved., Methods: To address this question we used 1) myeloid-cell-specific iNOS knockout mice in chronic smoke exposure and 2) co-cultures of macrophages and pulmonary artery smooth muscle cells (PASMCs) to decipher underlying signalling pathways., Results: Myeloid-cell-specific iNOS knockout prevented smoke-induced PH but not emphysema in mice. Moreover, iNOS deletion in myeloid cells ameliorated the increase in expression of CD206, a marker of M2 polarisation, on interstitial macrophages. Importantly, the observed effects on lung macrophages were hypoxia-independent, as these mice developed hypoxia-induced PH. In vitro , smoke-induced PASMC proliferation in co-cultures with M2-polarised macrophages could be abolished by iNOS deletion in phagocytic cells, as well as by extracellular signal-regulated kinase inhibition in PASMCs. Crucially, CD206-positive and iNOS-positive macrophages accumulated in proximity of remodelled vessels in the lungs of COPD patients, as shown by immunohistochemistry., Conclusion: In summary, our results demonstrate that iNOS deletion in myeloid cells confers protection against PH in smoke-exposed mice and provide evidence for an iNOS-dependent communication between M2-like macrophages and PASMCs in underlying pulmonary vascular remodelling., Competing Interests: Conflict of interest: M. Gredic has nothing to disclose. Conflict of interest: C-Y. Wu has nothing to disclose. Conflict of interest: S. Hadzic has nothing to disclose. Conflict of interest: O. Pak has nothing to disclose. Conflict of interest: R. Savai has nothing to disclose. Conflict of interest: B. Kojonazarov has nothing to disclose. Conflict of interest: S. Doswada has nothing to disclose. Conflict of interest: A. Weiss has nothing to disclose. Conflict of interest: A. Weigert has nothing to disclose. Conflict of interest: A. Guenther has nothing to disclose. Conflict of interest: R.P. Brandes has nothing to disclose. Conflict of interest: R.T. Schermuly has nothing to disclose. Conflict of interest: F. Grimminger has nothing to disclose. Conflict of interest: W. Seeger reports personal fees from Actelion, Bayer AG, Novartis, Vectura, Medspray and United Therapeutics, outside the submitted work. Conflict of interest: N. Sommer reports personal fees from Actelion, outside the submitted work. Conflict of interest: S. Kraut has nothing to disclose. Conflict of interest: N. Weissmann reports grants from the German Research Foundation, during the conduct of the study; and has a patent L-NIL as inhibitor for regenerating the lung of a patient suffering from COPD (EP2591777A2) issued., (Copyright ©The authors 2022.)

Aims: We sought to assess the feasibility of constructing right ventricular (RV) pressure-volume (PV) loops solely by echocardiography., Methods and Results: We performed RV conductance and pressure wire (PW) catheterization with simultaneous echocardiography in 35 patients with pulmonary hypertension. To generate echocardiographic PV loops, a reference RV pressure curve was constructed using pooled PW data from the first 20 patients (initial cohort). Individual pressure curves were then generated by adjusting the reference curve according to RV isovolumic and ejection phase duration and estimated RV systolic pressure. The pressure curves were synchronized with echocardiographic volume curves. We validated the reference curve in the remaining 15 patients (validation cohort). Methods were compared with correlation and Bland-Altman analysis. In the initial cohort, echocardiographic and conductance-derived PV loop parameters were significantly correlated {rho = 0.8053 [end-systolic elastance (Ees)], 0.8261 [Ees/arterial elastance (Ea)], and 0.697 (stroke work); all P < 0.001}, with low bias [-0.016 mmHg/mL (Ees), 0.1225 (Ees/Ea), and -39.0 mmHg mL (stroke work)] and acceptable limits of agreement. Echocardiographic and PW-derived Ees were also tightly correlated, with low bias (-0.009 mmHg/mL) and small limits of agreement. Echocardiographic and conductance-derived Ees, Ees/Ea, and stroke work were also tightly correlated in the validation cohort (rho = 0.9014, 0.9812, and 0.9491, respectively; all P < 0.001), with low bias (0.0173 mmHg/mL, 0.0153, and 255.1 mmHg mL, respectively) and acceptable limits., Conclusion: The novel echocardiographic method is an acceptable alternative to invasively measured PV loops to assess contractility, RV-arterial coupling, and RV myocardial work. Further validation is warranted., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions, please email: journals.permissions@oup.com.)

Background: Pulmonary hypertension (PH) is a life-threatening disease, characterized by excessive pulmonary vascular remodeling, leading to elevated pulmonary arterial pressure and right heart hypertrophy. PH can be caused by chronic hypoxia, leading to hyper-proliferation of pulmonary arterial smooth muscle cells (PASMCs) and apoptosis-resistant pulmonary microvascular endothelial cells (PMVECs). On reexposure to normoxia, chronic hypoxia-induced PH in mice is reversible. In this study, the authors aim to identify novel candidate genes involved in pulmonary vascular remodeling specifically in the pulmonary vasculature., Methods: After microarray analysis, the authors assessed the role of SPARC (secreted protein acidic and rich in cysteine) in PH using lung tissue from idiopathic pulmonary arterial hypertension (IPAH) patients, as well as from chronically hypoxic mice. In vitro studies were conducted in primary human PASMCs and PMVECs. In vivo function of SPARC was proven in chronic hypoxia-induced PH in mice by using an adeno-associated virus-mediated Sparc knockdown approach., Results: C57BL/6J mice were exposed to normoxia, chronic hypoxia, or chronic hypoxia with subsequent reexposure to normoxia for different time points. Microarray analysis of the pulmonary vascular compartment after laser microdissection identified Sparc as one of the genes downregulated at all reoxygenation time points investigated. Intriguingly, SPARC was vice versa upregulated in lungs during development of hypoxia-induced PH in mice as well as in IPAH, although SPARC plasma levels were not elevated in PH. TGF-β1 (transforming growth factor β1) or HIF2A (hypoxia-inducible factor 2A) signaling pathways induced SPARC expression in human PASMCs. In loss of function studies, SPARC silencing enhanced apoptosis and reduced proliferation. In gain of function studies, elevated SPARC levels induced PASMCs, but not PMVECs, proliferation. Coculture and conditioned medium experiments revealed that PMVECs-secreted SPARC acts as a paracrine factor triggering PASMCs proliferation. Contrary to the authors' expectations, in vivo congenital Sparc knockout mice were not protected from hypoxia-induced PH, most probably because of counter-regulatory proproliferative signaling. However, adeno-associated virus-mediated Sparc knockdown in adult mice significantly improved hemodynamic and cardiac function in PH mice., Conclusions: This study provides evidence for the involvement of SPARC in the pathogenesis of human PH and chronic hypoxia-induced PH in mice, most likely by affecting vascular cell function.

Rationale: The MSTs (mammalian Ste20-like kinases) 1/2 are members of the HIPPO pathway that act as growth suppressors in adult proliferative diseases. Pulmonary arterial hypertension (PAH) manifests by increased proliferation and survival of pulmonary vascular cells in small PAs, pulmonary vascular remodeling, and the rise of pulmonary arterial pressure. The role of MST1/2 in PAH is currently unknown., Objective: To investigate the roles and mechanisms of the action of MST1 and MST2 in PAH., Methods and Results: Using early-passage pulmonary vascular cells from PAH and nondiseased lungs and mice with smooth muscle-specific tamoxifen-inducible Mst1/2 knockdown, we found that, in contrast to canonical antiproliferative/proapoptotic roles, MST1/2 act as proproliferative/prosurvival molecules in human PAH pulmonary arterial vascular smooth muscle cells and pulmonary arterial adventitial fibroblasts and support established pulmonary vascular remodeling and pulmonary hypertension in mice with SU5416/hypoxia-induced pulmonary hypertension. By using unbiased proteomic analysis, gain- and loss-of function approaches, and pharmacological inhibition of MST1/2 kinase activity by XMU-MP-1, we next evaluated mechanisms of regulation and function of MST1/2 in PAH pulmonary vascular cells. We found that, in PAH pulmonary arterial adventitial fibroblasts, the proproliferative function of MST1/2 is caused by IL-6-dependent MST1/2 overexpression, which induces PSMC6-dependent downregulation of forkhead homeobox type O 3 and hyperproliferation. In PAH pulmonary arterial vascular smooth muscle cells, MST1/2 acted via forming a disease-specific interaction with BUB3 and supported ECM (extracellular matrix)- and USP10-dependent BUB3 accumulation, upregulation of Akt-mTORC1, cell proliferation, and survival. Supporting our in vitro observations, smooth muscle-specific Mst1/2 knockdown halted upregulation of Akt-mTORC1 in small muscular PAs of mice with SU5416/hypoxia-induced pulmonary hypertension., Conclusions: Together, this study describes a novel proproliferative/prosurvival role of MST1/2 in PAH pulmonary vasculature, provides a novel mechanistic link from MST1/2 via BUB3 and forkhead homeobox type O to the abnormal proliferation and survival of pulmonary arterial vascular smooth muscle cells and pulmonary arterial adventitial fibroblasts, remodeling and pulmonary hypertension, and suggests new target pathways for therapeutic intervention.

Background: γ-glutamyl transferase (GGT), the aspartate aminotransferase/alanine aminotransferase (AST/ALT) ratio, and the neutrophil-to-lymphocyte ratio (NLR) are prognostic biomarkers in several cardiovascular diseases, but their relevance in pulmonary hypertension (PH) is not fully understood. We aimed to assess their prognostic value in patients with pulmonary arterial hypertension (PAH) and chronic thromboembolic PH (CTEPH)., Methods: We retrospectively analyzed 731 incident patients with idiopathic PAH or CTEPH who entered the Giessen PH registry during 1993-2019. A risk stratification score based on GGT, AST/ALT ratio, and NLR tertiles was compared with a truncated version of the European Society of Cardiology/European Respiratory Society (ESC/ERS) risk stratification scheme. Associations with survival were evaluated using Kaplan-Meier and Cox regression analyses. External validation was performed in 311 patients with various types of PAH or CTEPH from a second German center., Results: GGT levels, AST/ALT, and NLR independently predicted mortality at baseline and during follow-up. The scoring system based on these biomarkers predicted mortality at baseline and during follow-up (both log-rank p < 0.001; hazard ratio [95% confidence interval], high vs low risk: baseline, 7.6 [3.9, 15.0]; follow-up, 13.3 [4.8, 37.1]). Five-year survival of low, intermediate, and high risk groups was 92%, 76%, and 51%, respectively, at baseline and 95%, 78%, and 50%, respectively, during follow-up. Our scoring system showed characteristics comparable to the ESC/ERS scheme, and predicted mortality in the validation cohort., Conclusion: GGT, AST/ALT, and NLR were reliable prognostic biomarkers at baseline and during follow-up, with predictive power comparable to the gold standard for risk stratification., Competing Interests: Disclosure statement Mr Yogeswaran, Dr Tello, Mr Lund, Dr Klose, Dr Harbaum, Dr Sommer, Dr Oqueka, Dr Hennigs, and Dr Grimminger have declared no conflict of interest. Dr Seeger has received speaker/consultancy fees/fees for participation in advisory boards from Pfizer, Novartis, United Therapeutics, Actelion, Vectura, Savara, Medspray, and Bayer Pharma AG. Dr Ghofrani has received consultancy fees from Bayer, Actelion, Pfizer, Merck, GSK, and Novartis; fees for participation in advisory boards from Bayer, Pfizer, GSK, Actelion, and Takeda; lecture fees from Bayer HealthCare, GSK, Actelion, and Encysive/Pfizer; industry-sponsored grants from Bayer HealthCare, Aires, Encysive/Pfizer, and Novartis; and sponsored grants from the German Research Foundation, Excellence Cluster Cardiopulmonary Research, and the German Ministry for Education and Research. Dr Richter has received support from United Therapeutics and Bayer Pharma AG, and speaker fees from Actelion, Mundipharma, Roche, and OMT. Dr Gall has received fees from Actelion, AstraZeneca, Bayer, BMS, GSK, Janssen-Cilag, Lilly, MSD, Novartis, OMT, Pfizer, and United Therapeutics., (Copyright © 2021 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Background: Patients with pulmonary hypertension (PH) frequently show preserved right ventricular (RV) function at rest. However, volume challenge may uncover pending RV dysfunction. We aimed to assess the physiological and prognostic impact of RV-pulmonary arterial (RV-PA) uncoupling during volume challenge in patients with precapillary PH., Methods: We prospectively assessed 32 patients with PH (pulmonary arterial hypertension, n = 27; inoperable chronic thromboembolic disease, n = 5) and 4 controls using invasive pressure-volume (PV) catheterization. PV loops were recorded during preload reduction (balloon occlusion of inferior vena cava; baseline) and acute volume loading (200 ml saline in 20 s). Contractility (multi-beat end-systolic elastance [Ees]), arterial elastance (Ea), and RV-PA coupling (Ees/Ea) were obtained at baseline and at maximum volume loading (MVL)., Results: Median [interquartile range] time to MVL was 19 [18-22] s. Ees/Ea significantly declined from baseline (0.89 [0.69-1.23]) to MVL (0.16 [0.12-0.34]; p < 0.001) in patients with PH but remained stable in controls (baseline: 1.08 [0.94-1.80]; MVL: 1.01 [0.80-2.49]; p = 0.715). The same pattern was observed for Ees, while Ea remained unchanged. The percent decline of RV-PA coupling (ΔEes/Ea) during fluid challenge was significantly associated with pulmonary resting hemodynamics, RV ejection fraction (RVEF), and RV end-diastolic volume. Kaplan-Meier analysis revealed that patients with PH who had a smaller ΔEes/Ea (<-65%) had a significantly better prognosis (log-rank p = 0.0389). In multivariate Cox regression analysis, clinical worsening was predicted by ΔEes/Ea (hazard ratio: 0.96 [95% confidence interval: 0.93-1.00]) and RVEF (hazard ratio: 0.95 [95% confidence interval: 0.92-0.98])., Conclusions: Assessment of PV loops during fluid challenge uncovers exhausted RV coupling reserve with severely reduced contractility in PH. RV-PA uncoupling during volume challenge can be predicted by pulmonary resting hemodynamics and RVEF. RV-PA uncoupling during fluid challenge and RVEF (as a noninvasive correlate) are predictors of clinical worsening., Clinical Trial Registration: URL: https://www.clinicaltrials.gov. Unique identifier: NCT03403868 (January 19, 2018)., (Copyright © 2021 International Society for Heart and Lung Transplantation. All rights reserved.)

Purpose: The aim of this study was to assess effects of balloon pulmonary angioplasty (BPA) on right ventricular (RV) mechanical dysfunction in patients with inoperable chronic thromboembolic pulmonary hypertension (CTEPH) via MRI., Method: MRI at 1.5 Tesla and right heart catheterization were performed before and 6 months after BPA in 30 CTEPH patients (mean age 63.4 ± 10.6 years; 17 female). Feature-tracking strain analysis, including global longitudinal (GLS), circumferential (GCS), and radial (GRS) strain, was performed and compared with right ventricular function, myocardial remodelling (assessed by native T1 times), and pulmonary haemodynamics (mean pulmonary arterial pressure and pulmonary vascular resistance)., Results: RVEF (35.9% to 48.4%) increased and mPAP (42.1 mmHg to 33.1 mmHg) and PVR (551.8 to 377.7 dyn∙s/cm 5 ) decreased after BPA (all p < 0.0001). Moreover, RV strain increased (GLS -19.9 to -24.0%, p = 0.0003; GCS -9.4 to -11.0%, p = 0.0022; GRS 38.2 to 50.7%, p = 0.001) and septal native area-adjusted T1 time (AA-T1) decreased (1019.4 to 988.7 ms, p < 0.0001). GLS revealed the best correlations with RVEF (before BPA r = -0.75; after BPA r = -0.54), mPAP (r = 0.36; r = 0.52), PVR (r = 0.49; r = 0.48), and AA-T1 (r = 0.44; 0.19)., Conclusion: RV mechanical dysfunction, pulmonary haemodynamics, and myocardial remodelling are markedly improved by BPA. Moreover, RV strain values showed good correlations with RV function, pulmonary haemodynamics, and myocardial remodelling. Therefore, strain analysis might provide new insights regarding therapy outcome, monitoring, and prognosis., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Aims: The pulmonary vascular tone and hypoxia-induced alterations of the pulmonary vasculature may be regulated by the mitochondrial membrane permeability transition pore (mPTP) that controls mitochondrial calcium load and apoptosis. We thus investigated, if the mitochondrial proteins p66shc and cyclophilin D (CypD) that regulate mPTP opening affect the pulmonary vascular tone., Methods and Results: Mice deficient for p66shc (p66shc-/-), CypD (CypD-/-), or both proteins (p66shc/CypD-/-) exhibited decreased pulmonary vascular resistance (PVR) compared to wild-type mice determined in isolated lungs and in vivo. In contrast, systemic arterial pressure was only lower in CypD-/- mice. As cardiac function and pulmonary vascular remodelling did not differ between genotypes, we determined alterations of vascular contractility in isolated lungs and calcium handling in pulmonary arterial smooth muscle cells (PASMC) as underlying reason for decreased PVR. Potassium chloride (KCl)-induced pulmonary vasoconstriction and KCl-induced cytosolic calcium increase determined by Fura-2 were attenuated in all gene-deficient mice. In contrast, KCl-induced mitochondrial calcium increase determined by the genetically encoded Mito-Car-GECO and calcium retention capacity were increased only in CypD-/- and p66shc/CypD-/- mitochondria indicating that decreased mPTP opening affected KCl-induced intracellular calcium peaks in these cells. All mouse strains showed a similar pulmonary vascular response to chronic hypoxia, while acute hypoxic pulmonary vasoconstriction was decreased in gene-deficient mice indicating that CypD and p66shc regulate vascular contractility but not remodelling., Conclusions: We conclude that p66shc specifically regulates the pulmonary vascular tone, while CypD also affects systemic pressure. However, only CypD acts via regulation of mPTP opening and mitochondrial calcium regulation., (© The Author(s) 2020. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Purpose:  The purpose of this study was to assess the diagnostic accuracy of computed tomography pulmonary angiogram (CTPA) including dual energy and reconstruction of iodine maps for diagnosing CTEPH. This method for detecting embolisms and perfusion failures was compared with V/Q-SPECT. An additional purpose was to compare the applied radiation dose of both techniques., Materials and Methods:  71 patients (49 women) with suspected CTEPH were included in this prospective study. The patients received a V/Q-SPECT and a dual-energy CTPA. Iodine maps were reconstructed from the data set. CTPA and the iodine maps were read by an experienced radiologist unaware of the clinical information as well as the results of the V/Q-SPECT. Results were compared to the V/Q-SPECT. DLP and the applied amount of radionuclides (MAA, Technegas) were obtained for comparison of radiation dose., Results:  For the diagnosis of CTEPH, the sensitivity of DECT was 1.000, specificity 0.966, PPV 0.867 and NPV 1.000, respectively. There was not a considerable difference in the x-ray exposure between the DECT examination and the V/Q-SPECT (1.892 mSv vs. 1.911 mSv; p = 0.6115). Both examination modalities were highly consistent regarding the classification of pathological segments (1177/1278 segments, 92,09 %, κ = 0,5938)., Conclusion:  This study presents the DECT, in combination with reconstructed iodine maps, as a potential alternative to the current imaging technique of first choice, V/Q-SPECT. For creating future prospective diagnostic algorithms, the implementation of DECT screening with iodine maps should be considered., Key Points: · DECT correctly identified all CTEPH patients.. · There is substantial agreement between DECT and V/Q-SPECT in the classification of pathological segments.. · There is no significant difference in radiation exposure during DECT examination and V/Q-SPECT examination.. · Reduced radiation dose does not negatively impact image quality.., Citation Format: · Schüßler A, Richter M, Tello K et al. Evaluation der diagnostischen Genauigkeit und der Strahlendosis der Dual-Energy-Computertomografie (DECT) bei chronisch thromboembolischer pulmonaler Hypertonie (CTEPH). Fortschr Röntgenstr 2021; 193: 1318 - 1326., Competing Interests: The authors declare that they have no conflict of interest., (Thieme. All rights reserved.)

Vascular remodeling is a prominent feature of pulmonary hypertension. This process involves increased muscularization of already muscularized vessels as well as neo-muscularization of non-muscularized vessels. The cell-of-origin of the newly formed vascular smooth muscle cells has been a subject of intense debate in recent years. Identifying these cells may have important clinical implications since it opens the door for attempts to therapeutically target the progenitor cells and/or reverse the differentiation of their progeny. In this context, the dominant model is that these cells derive from pre-existing smooth muscle cells that are activated in response to injury. In this mini review, we present the evidence that is in favor of this model and, at the same time, highlight other studies indicating that there are alternative cellular sources of vascular smooth muscle cells in pulmonary vascular remodeling.

Aim: This study assessed the utility of osteopontin (OPN) and galectin-3 (Gal-3) as biomarkers of maladaptive right ventricular remodeling in pulmonary hypertension (PH). Materials & methods: We examined plasma levels of OPN and Gal-3 in patients with PH (n = 62), dilated cardiomyopathy (n = 34), left ventricular hypertrophy (LVH; n = 47), and controls without right ventricle (RV) or LV abnormalities (n = 38). Results: OPN and Gal-3 levels were higher in PH, dilated cardiomyopathy and LVH than in the controls. OPN concentrations in PH patients with maladaptive RV were significantly higher than in those with adaptive RV. Gal-3 did not differentiate between adaptive and maladaptive RV remodeling in PH. OPN and Gal-3 levels did not correlate with parameters of LV remodeling. Conclusion: OPN is a potential biomarker of RV maladaptation.

Right ventricular (RV) function determines outcome in pulmonary arterial hypertension (PAH). RV pressure-volume loops, the gold standard for measuring RV function, are difficult to analyze. Our aim was to investigate whether simple assessments of RV pressure-volume loop morphology and RV systolic pressure differential reflect PAH severity and RV function. We analyzed multibeat RV pressure-volume loops (obtained by conductance catheterization with preload reduction) in 77 patients with PAH and 15 patients without pulmonary hypertension in two centers. Patients were categorized according to their pressure-volume loop shape (triangular, quadratic, trapezoid, or notched). RV systolic pressure differential was defined as end-systolic minus beginning-systolic pressure (ESP - BSP), augmentation index as ESP - BSP/pulse pressure, pulmonary arterial capacitance (PAC) as stroke volume/pulse pressure, and RV-arterial coupling as end-systolic/arterial elastance (Ees/Ea). Trapezoid and notched pressure-volume loops were associated with the highest afterload (Ea), augmentation index, pulmonary vascular resistance (PVR), mean pulmonary arterial pressure, stroke work, B-type natriuretic peptide, and the lowest Ees/Ea and PAC. Multivariate linear regression identified Ea, PVR, and stroke work as the main determinants of ESP - BSP. ESP - BSP also significantly correlated with multibeat Ees/Ea (Spearman's ρ: -0.518, P < 0.001). A separate retrospective analysis of 113 patients with PAH showed that ESP - BSP obtained by routine right heart catheterization significantly correlated with a noninvasive surrogate of RV-arterial coupling (tricuspid annular plane systolic excursion/pulmonary arterial systolic pressure ratio; ρ: -0.376, P < 0.001). In conclusion, pressure-volume loop shape and RV systolic pressure differential predominately depend on afterload and PAH severity and reflect RV-arterial coupling in PAH.

Chronic obstructive pulmonary disease (COPD) is a major cause of death and a still incurable disease, comprising emphysema and chronic bronchitis. In addition to airflow limitation, patients with COPD can suffer from pulmonary hypertension (PH). Doxycycline, an antibiotic from the tetracycline family, in addition to its pronounced antimicrobial activity, acts as a matrix metalloproteinase (MMP) inhibitor and has anti-inflammatory properties. Furthermore, doxycycline treatment exhibited a beneficial effect in several preclinical cardiovascular disease models. In preclinical research, doxycycline is frequently employed for gene expression modulation in Tet-On/Tet-Off transgenic animal models. Therefore, it is crucial to know whether doxycycline treatment in Tet-On/Tet-Off systems has effects independent of gene expression modulation by such systems. Against this background, we assessed the possible curative effects of long-term doxycycline administration in a mouse model of chronic CS exposure. Animals were exposed to cigarette smoke (CS) for 8 mo and then subsequently treated with doxycycline for additional 3 mo in room air conditions. Doxycycline decreased the expression of MMPs and general pro-inflammatory markers in the lungs from CS-exposed mice. This downregulation was, however, insufficient to ameliorate CS-induced emphysema or PH. Tet-On/Tet-Off induction by doxycycline in such models is a feasible genetic approach to study curative effects at least in established CS-induced emphysema and PH. However, we report several parameters that are influenced by doxycycline and use of a Tet-On/Tet-Off system when evaluating those parameters should be interpreted with caution.

The aim of our study was to analyse the protein expression of cartilage intermediate layer protein (CILP)1 in a mouse model of right ventricular (RV) pressure overload and to evaluate CILP1 as a biomarker of cardiac remodelling and maladaptive RV function in patients with pulmonary hypertension (PH).Pulmonary artery banding was performed in 14 mice; another nine mice underwent sham surgery. CILP1 protein expression was analysed in all hearts using Western blotting and immunostaining. CILP1 serum concentrations were measured in 161 patients (97 with adaptive and maladaptive RV pressure overload caused by PH; 25 with left ventricular (LV) hypertrophy; 20 with dilative cardiomyopathy (DCM); 19 controls without LV or RV abnormalities)In mice, the amount of RV CILP1 was markedly higher after banding than after sham. Control patients had lower CILP1 serum levels than all other groups (p<0.001). CILP1 concentrations were higher in PH patients with maladaptive RV function than those with adaptive RV function (p<0.001), LV pressure overload (p<0.001) and DCM (p=0.003). CILP1 showed good predictive power for maladaptive RV in receiver operating characteristic analysis (area under the curve (AUC) 0.79). There was no significant difference between the AUCs of CILP1 and N-terminal pro-brain natriuretic peptide (NT-proBNP) (AUC 0.82). High CILP1 (cut-off value for maladaptive RV of ≥4373 pg·mL -1 ) was associated with lower tricuspid annular plane excursion/pulmonary artery systolic pressure ratios (p<0.001) and higher NT-proBNP levels (p<0.001).CILP1 is a novel biomarker of RV and LV pathological remodelling that is associated with RV maladaptation and ventriculoarterial uncoupling in patients with PH., Competing Interests: Conflict of interest: S. Keranov reports grants from German Research Foundation (Project B07, Collaborative Research Center 1213-Pulmonary Hypertension and Cor Pulmonale), during the conduct of the study. Conflict of interest: O. Dörr reports grants from German Research Foundation (Project B07, Collaborative Research Center 1213-Pulmonary Hypertension and Cor Pulmonale), during the conduct of the study. Conflict of interest: L. Jafari has nothing to disclose. Conflict of interest: C. Troidl has nothing to disclose. Conflict of interest: C. Liebetrau reports grants from German Research Foundation (Project B07, Collaborative Research Center 1213-Pulmonary Hypertension and Cor Pulmonale), during the conduct of the study. Conflict of interest: S. Kriechbaum reports grants from German Research Foundation (Project B07, Collaborative Research Center 1213-Pulmonary Hypertension and Cor Pulmonale), during the conduct of the study. Conflict of interest: T. Keller has nothing to disclose. Conflict of interest: S. Voss has nothing to disclose. Conflict of interest: T. Bauer has nothing to disclose. Conflict of interest: J. Lorenz has nothing to disclose. Conflict of interest: M.J. Richter has nothing to disclose. Conflict of interest: K. Tello has nothing to disclose. Conflict of interest: H. Gall has nothing to disclose. Conflict of interest: H.A. Ghofrani has nothing to disclose. Conflict of interest: E. Mayer has nothing to disclose. Conflict of interest: C.B. Wiedenroth has nothing to disclose. Conflict of interest: S. Guth has nothing to disclose. Conflict of interest: H. Lörchner has nothing to disclose. Conflict of interest: J. Pöling has nothing to disclose. Conflict of interest: P. Chelladurai has nothing to disclose. Conflict of interest: S.S. Pullamsetti has nothing to disclose. Conflict of interest: T. Braun has nothing to disclose. Conflict of interest: W. Seeger has nothing to disclose. Conflict of interest: C.W. Hamm reports grants from German Research Foundation (Project B07, Collaborative Research Center 1213-Pulmonary Hypertension and Cor Pulmonale), during the conduct of the study. Conflict of interest: H. Nef reports grants from German Research Foundation (Project B07, Collaborative Research Center 1213-Pulmonary Hypertension and Cor Pulmonale), during the conduct of the study., (Copyright ©ERS 2021.)

Pulmonary hypertension (PH) is characterized by a progressive elevation of mean arterial pressure followed by right ventricular failure and death. Previous studies have indicated that numerous inhibitors of receptor tyrosine kinase signaling could be either beneficial or detrimental for the treatment of PH. Here we investigated the therapeutic potential of the multi-kinase inhibitor regorafenib (BAY 73-4506) for the treatment of PH. A peptide-based kinase activity assay was performed using the PamStation ® 12 platform. The 5-bromo-2'-deoxyuridine proliferation and transwell migration assays were utilized in pulmonary arterial smooth muscle cells (PASMCs). Regorafenib was administered to monocrotaline- and hypoxia-induced PH in rats and mice, respectively. Functional parameters were analyzed by hemodynamic and echocardiographic measurements. The kinase activity assay revealed upregulation of twenty-nine kinases in PASMCs from patients with idiopathic PAH (IPAH), of which fifteen were established as potential targets of regorafenib. Regorafenib showed strong anti-proliferative and anti-migratory effects in IPAH-PASMCs compared to the control PASMCs. Both experimental models indicated improved cardiac function and reduced pulmonary vascular remodeling upon regorafenib treatment. In lungs from monocrotaline (MCT) rats, regorafenib reduced the phosphorylation of c-Jun N-terminal kinase and extracellular signal-regulated kinase 1/2. Overall, our data indicated that regorafenib plays a beneficial role in experimental PH., Competing Interests: The authors declare no conflict of interest.

Pulmonary hypertension (PH) is a progressive complex fatal disease of multiple etiologies. Hyperproliferation and resistance to apoptosis of vascular cells of intimal, medial, and adventitial layers of pulmonary vessels trigger excessive pulmonary vascular remodeling and vasoconstriction in the course of pulmonary arterial hypertension (PAH), a subgroup of PH. Multiple gene mutation/s or dysregulated gene expression contribute to the pathogenesis of PAH by endorsing the proliferation and promoting the resistance to apoptosis of pulmonary vascular cells. Given the vital role of these cells in PAH progression, the development of safe and efficient-gene therapeutic approaches that lead to restoration or down-regulation of gene expression, generally involved in the etiology of the disease is the need of the hour. Currently, none of the FDA-approved drugs provides a cure against PH, hence innovative tools may offer a novel treatment paradigm for this progressive and lethal disorder by silencing pathological genes, expressing therapeutic proteins, or through gene-editing applications. Here, we review the effectiveness and limitations of the presently available gene therapy approaches for PH. We provide a brief survey of commonly existing and currently applicable gene transfer methods for pulmonary vascular cells in vitro and describe some more recent developments for gene delivery existing in the field of PH in vivo.

Background and Purpose: Chronic obstructive pulmonary disease, encompassing chronic airway obstruction and lung emphysema, is a major worldwide health problem and a severe socio-economic burden. Evidence previously provided by our group has shown that inhibition of inducible NOS (iNOS) prevents development of mild emphysema in a mouse model of chronic tobacco smoke exposure and can even trigger lung regeneration. Moreover, we could demonstrate that pulmonary hypertension is not only abolished in cigarette smoke-exposed iNOS -/- mice but also precedes emphysema development. Possible regenerative effects of pharmacological iNOS inhibition in more severe models of emphysema not dependent on tobacco smoke, however, are hitherto unknown., Experimental Approach: We have established a mouse model using a single dose of porcine pancreatic elastase or saline, intratracheally instilled in C57BL/6J mice. Emphysema, as well as pulmonary hypertension development was determined by both structural and functional measurements., Key Results: Our data revealed that (i) emphysema is fully established after 21 days, with the same degree of emphysema after 21 and 28 days post instillation, (ii) emphysema is stable for at least 12 weeks and (iii) pulmonary hypertension is evident, in contrast to smoke models, only after emphysema development. Oral treatment with the iNOS inhibitor N(6)-(1-iminoethyl)-l-lysine (L-NIL) was started after emphysema establishment and continued for 12 weeks. This resulted in significant lung regeneration, evident in the improvement of emphysema and reversal of pulmonary hypertension., Conclusion and Implications: Our data indicate that iNOS is a potential new therapeutic target to treat severe emphysema and associated pulmonary hypertension., Linked Articles: This article is part of a themed issue on Risk factors, comorbidities, and comedications in cardioprotection. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.1/issuetoc., (© 2020 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Epigenetic mechanisms, including DNA methylation and histone post-translational modifications (PTMs), have been known to regulate chromatin structure and lineage-specific gene expression during cardiovascular development and disease. However, alterations in the landscape of histone PTMs and their contribution to the pathogenesis of incurable cardiovascular diseases such as pulmonary hypertension (PH) and associated right heart failure (RHF) remain largely unexplored. This review focusses on the studies in PH and RHF that investigated the gene families that write (histone acetyltransferases), read (bromodomain-containing proteins) or erase (histone deacetylases [HDACs] and sirtuins [SIRT]) acetyl moieties from the ε-amino group of lysine residues of histones and non-histone proteins. Analysis of cells and tissues isolated from the in vivo preclinical models of PH and human pulmonary arterial hypertension not only confirmed significant alterations in the expression levels of multiple HDACs, SIRT1, SIRT3 and BRD4 proteins but also demonstrated their strong association to proliferative, inflammatory and fibrotic phenotypes linked to the pathological vascular remodelling process. Due to the reversible nature of post-translational protein acetylation, the therapeutic efficacy of numerous small-molecule inhibitors (vorinostat, valproic acid, sodium butyrate, mocetinostat, entinostat, tubastatin A, apabetalone, JQ1 and resveratrol) have been evaluated in different preclinical models of cardiovascular disease, which revealed the promising therapeutic benefits of targeting histone acetylation pathways in the attenuation of cardiac hypertrophy, fibrosis, left heart dysfunction, PH and RHF. This review also emphasizes the need for deeper molecular insights into the contribution of epigenetic changes to PH pathogenesis and therapeutic evaluation of isoform-specific modulation in ex vivo and in vivo models of PH and RHF. LINKED ARTICLES: This article is part of a themed issue on Risk factors, comorbidities, and comedications in cardioprotection. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.1/issuetoc., (© 2019 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

In pulmonary arterial hypertension (PAH), progressive structural remodeling accounts for the pulmonary vasculopathy including the obliteration of the lung vasculature that causes an increase in vascular resistance and mean blood pressure in the pulmonary arteries ultimately leading to right heart failure-mediated death. Deciphering the molecular details of aberrant signaling of pulmonary vascular cells in PAH is fundamental for the development of new therapeutic strategies. We aimed to identify kinases as new potential drug targets that are dysregulated in PAH by means of a peptide-based kinase activity assay. We performed a tyrosine kinase-dependent phosphorylation assay using 144 selected microarrayed substrate peptides. The differential signature of phosphopeptides was used to predict alterations in tyrosine kinase activities in human pulmonary arterial smooth muscle cells (HPASMCs) from patients with idiopathic PAH (IPAH) compared with healthy control cells. Thereby, we observed an overactivation and an increased expression of Jak2 (Janus kinase 2) in HPASMCs from patients with IPAH as compared with controls. In vitro , IL-6-induced proliferation and migration of HPASMCs from healthy individuals as well as from patients with IPAH were reduced in a dose-dependent manner by the U.S. Food and Drug Administration-approved Jak1 and Jak2 inhibitor ruxolitinib. In vivo , ruxolitinib therapy in two experimental models of pulmonary arterial hypertension dose-dependently attenuated the elevation in pulmonary arterial pressure, partially reduced right ventricular hypertrophy, and almost completely restored cardiac index without signs of adverse events on cardiac function. Therefore, we propose that ruxolitinib may present a novel therapeutic option for patients with PAH by reducing pulmonary vascular remodeling through effectively blocking Jak2-Stat3 (signal transducer of activators of transcription)-mediated signaling pathways.

Adaptation of right ventricular (RV) function to increased afterload-known as RV-arterial coupling-is a key determinant of prognosis in pulmonary hypertension. However, measurement of RV-arterial coupling is a complex, invasive process involving analysis of the RV pressure-volume relationship during preload reduction over multiple cardiac cycles. Simplified methods have therefore been proposed, including echocardiographic and cardiac MRI approaches. This review describes the available methods for assessment of RV function and RV-arterial coupling and the effects of pharmacotherapy on these variables. Overall, pharmacotherapies for pulmonary hypertension have shown beneficial effects on various measures of RV function, but it is often unclear if these are direct RV effects or indirect results of afterload reduction. Studies of the effects of pharmacotherapies on RV-arterial coupling are limited and mostly restricted to experimental models. Simplified methods to assess RV-arterial coupling should be validated and incorporated into routine clinical follow-up and future clinical trials. LINKED ARTICLES: This article is part of a themed issue on Risk factors, comorbidities, and comedications in cardioprotection. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.1/issuetoc., (© 2019 The British Pharmacological Society.)

Rationale: Long noncoding RNAs (lncRNAs) are emerging as important regulators of diverse biological functions. Their role in pulmonary arterial hypertension (PAH) remains to be explored. Objectives: To elucidate the role of TYKRIL (tyrosine kinase receptor-inducing lncRNA) as a regulator of p53/ PDGFRβ (platelet-derived growth factor receptor β) signaling pathway and to investigate its role in PAH. Methods: Pericytes and pulmonary arterial smooth muscle cells exposed to hypoxia and derived from patients with idiopathic PAH were analyzed with RNA sequencing. TYKRIL knockdown was performed in above-mentioned human primary cells and in precision-cut lung slices derived from patients with PAH. Measurements and Main Results: Using RNA sequencing data, TYKRIL was identified to be consistently upregulated in pericytes and pulmonary arterial smooth muscles cells exposed to hypoxia and derived from patients with idiopathic PAH. TYKRIL knockdown reversed the proproliferative ( n  = 3) and antiapoptotic ( n  = 3) phenotype induced under hypoxic and idiopathic PAH conditions. Owing to the poor species conservation of TYKRIL, ex vivo studies were performed in precision-cut lung slices from patients with PAH. Knockdown of TYKRIL in precision-cut lung slices decreased the vascular remodeling ( n  = 5). The number of proliferating cell nuclear antigen-positive cells in the vessels was decreased and the number of terminal deoxynucleotide transferase-mediated dUTP nick end label-positive cells in the vessels was increased in the LNA (locked nucleic acid)-treated group compared with control. Expression of PDGFRβ, a key player in PAH, was found to strongly correlate with TYKRIL expression in the patient samples ( n  = 12), and TYKRIL knockdown decreased PDGFRβ expression ( n  = 3). From the transcription factor-screening array, it was observed that TYKRIL knockdown increased the p53 activity, a known repressor of PDGFRβ. RNA immunoprecipitation using various p53 mutants demonstrated that TYKRIL binds to the N-terminal of p53 (an important region for p300 interaction with p53). The proximity ligation assay revealed that TYKRIL interferes with the p53-p300 interaction ( n  = 3) and regulates p53 nuclear translocation. Conclusions: TYKRIL plays an important role in PAH by regulating the p53/PDGFRβ axis.

Pulmonary hypertension (PH) is characterized by pulmonary artery remodeling that can subsequently culminate in right heart failure and premature death. Emerging evidence suggests that hypoxia-inducible factor (HIF) signaling plays a fundamental and pivotal role in the pathogenesis of PH. This Review summarizes the regulation of HIF isoforms and their impact in various PH subtypes, as well as the elaborate conditional and cell-specific knockout mouse studies that brought the role of this pathway to light. We also discuss the current preclinical status of pan- and isoform-selective HIF inhibitors, and propose new research areas that may facilitate HIF isoform-specific inhibition as a novel therapeutic strategy for PH and right heart failure.

PKGs are serine/threonine kinases. PKG1 has two isoforms-PKG1α and β. Inositol trisphosphate receptor (IP 3 R)-associated cGMP-kinase substrate 1 (IRAG1) is a substrate for PKG1β. IRAG1 is also known to further interact with IP 3 RI, which mediates intracellular Ca 2+ release. However, the role of IRAG1 in PH is not known. Herein, WT and IRAG1 KO mice were kept under normoxic or hypoxic (10% O 2 ) conditions for five weeks. Animals were evaluated for echocardiographic variables and went through right heart catheterization. Animals were further sacrificed to prepare lungs and right ventricular (RV) for immunostaining, western blotting, and pulmonary artery smooth muscle cell (PASMC) isolation. IRAG1 is expressed in PASMCs and downregulated under hypoxic conditions. Genetic deletion of IRAG1 leads to RV hypertrophy, increase in RV systolic pressure, and RV dysfunction in mice. Absence of IRAG1 in lung and RV have direct impacts on PKG1β expression. Attenuated PKG1β expression in IRAG1 KO mice further dysregulates other downstream candidates of PKG1β in RV. IRAG1 KO mice develop PH spontaneously. Our results indicate that PKG1β signaling via IRAG1 is essential for the homeostasis of PASMCs and RV. Disturbing this signaling complex by deleting IRAG1 can lead to RV dysfunction and development of PH in mice.

Background: The updated hemodynamic definition of pulmonary hypertension (PH) due to interstitial lung disease (ILD) differentiates severe and non-severe phenotypes, but no further risk stratification strategy has been established or validated for severe PH due to ILD. We aimed to assess the prognostic value of a truncated version of the European Society of Cardiology/European Respiratory Society (ESC/ERS) PH risk stratification scheme in severe PH due to ILD., Methods: We retrospectively analyzed 185 patients with severe PH (mean pulmonary artery pressure of ≥35 mm Hg or ≥25 mm Hg with cardiac index <2.0 liter/min/m 2 ) due to ILD who were enrolled in the Giessen PH Registry after being referred for invasive diagnostic work-up of suspected PH during 1995‒2018. A truncated ESC/ERS risk stratification scheme (based on 8 parameters from the full scheme) was applied. Kaplan-Meier and univariate Cox regression analyses were used to evaluate transplant-free survival and hazard ratios, respectively., Results: During follow-up (median [interquartile range]: 19 [7-40] months), 146 events occurred. Using baseline data for risk stratification, 5-year transplant-free survival of low-, intermediate-, and high-risk groups was 43%, 15%, and 4%, respectively (log-rank p = 0.010; hazard ratio of high- vs low-risk group: 3.116 [95% CI: 1.428-6.800]). Using follow-up data (at 11 [6.0-32.5] months) for risk stratification, 5-year survival of low-, intermediate-, and high-risk groups was 22%, 3%, and 0%, respectively (log-rank p = 0.005)., Conclusions: The truncated ESC/ERS scheme was clinically useful and demonstrated prognostic relevance in severe PH due to ILD., (Copyright © 2020. Published by Elsevier Inc.)

Pharmacological modulation of class I histone deacetylases (HDAC) has been evaluated as a therapeutic strategy for pulmonary hypertension (PH) in experimental models of PH. However, information of their expression, regulation and transcriptional targets in human PH and the therapeutic potential of isoform-selective enzyme modulation are lacking. Comprehensive analysis of expression and regulation of class I HDACs (HDAC1, HDAC2, HDAC3 and HDAC8) was performed in cardiopulmonary tissues and adventitial fibroblasts isolated from pulmonary arteries (PAAF) of idiopathic pulmonary arterial hypertension (IPAH) patients and healthy donors. Cellular functions and transcriptional targets of HDAC enzymes were investigated. Therapeutic effects of pan-HDAC (Vorinostat), class-selective (VPA) and isoform-selective (CAY10398, Romidepsin, PCI34051) HDAC inhibitors were evaluated ex vivo (IPAH-PAAF, IPAH-PASMC) and in vivo (rat chronic hypoxia-induced PH and zebrafish angiogenesis). Our screening identifies dysregulation of class I HDAC isoforms in IPAH. Particularly, HDAC1 and HDAC8 were consistently increased in IPAH-PAs and IPAH-PAAFs, whereas HDAC2 and HDAC8 showed predominant localization with ACTA2-expressing cells in extensively remodeled IPAH-PAs. Hypoxia not only significantly modulated protein levels of deacetylase (HDAC8), but also significantly caused dynamic changes in the global histone lysine acetylation levels (H3K4ac, H3K9/K14ac and H3K27ac). Importantly, isoform-specific RNA-interference revealed that HDAC isoforms regulate distinct subset of transcriptome in IPAH-PAAFs. Reduced transcript levels of KLF2 in IPAH-PAAFs was augmented by HDAC8 siRNA and HDAC inhibitors, which also attenuated IPAH-associated hyperproliferation and apoptosis-resistance ex vivo, and mitigated chronic hypoxia-induced established PH in vivo, at variable degree. Class I HDAC isoforms are significantly dysregulated in human PAH. Isoform-selective HDAC inhibition is a viable approach to circumvent off-target effects.

Background and Purpose: Recruitment and involvement of bone-/blood-derived circulating fibrocytes (CF) in the promotion of fibrotic tissue remodelling processes have been shown. However, their direct contribution to pathological changes is not clear. The present study investigates the causal role of CF in the pathogenesis of pulmonary hypertension (PH)., Experimental Approach: For selective ablation of CF, we applied the suicidal gene strategy with herpes simplex virus thymidine kinase (HSV-TK) and ganciclovir. The transgenic mice were generated, having HSV-TK-GFP transgene under the collagen 1 promoter. To selectively target CF, HSV-TK-GFP + bone marrow transplanted into irradiated wild type mice. These chimera mice were subjected to hypoxia for PH induction and ganciclovir for CF ablation., Key Results: In vivo CF ablation reduced right ventricular hypertrophy and vascular remodelling with reduced total collagen content. We quantified the CF recruited in the perivascular area and arterial wall of small pulmonary arteries. There was significant recruitment of CF in the lung in response to hypoxia. The characterization of CF showed the expression of CD45 and collagen1 (GFP) along with α-smooth muscle actin (αSMA)., Conclusion and Implications: Our data demonstrated that CF ablation has a potential impact on right ventricular hypertrophy and vascular remodelling in the setting of experimental pulmonary hypertension induced by hypoxia. The beneficial effects may be related to the direct contribution of fibrocytes or its paracrine effect on other resident cell types. Thus, clinical manipulation of CF may represent a novel therapeutic approach to ameliorate the disease state in pulmonary hypertension., (© 2020 The British Pharmacological Society.)

Aim: This study assessed the utility of SPARC-like protein 1 (SPARCL1) as a biomarker of maladaptive right ventricular (RV) function in patients with pulmonary hypertension (PH). Methods: In this prospective study, we examined SPARCL1 levels in 105 patients with adaptive ( n  = 34) and maladaptive RV ( n  = 32) pressure overload caused by PH, dilated cardiomyopathy (DCM, n  = 18) with LVEF < 35% and preserved RV function and controls without LV or RV abnormalities ( n  = 21). Results: The median SPARCL1 concentration in patients with maladaptive RV function was higher than in those with adaptive RV function ( p  < 0.01), DCM ( p  < 0.001) or controls ( p  < 0.001). Patients with adaptive RV function had higher SPARCL1 concentrations than controls ( p  < 0.05), whereas there was no difference between adaptive RV and DCM. SPARCL1 showed good predictive power for maladaptive RV (AUC 0.77, p  < 0.001) with an optimal cut-off value of 9.66 ng/ml. The TAPSE/PASP ratio was the only independent predictor of SPARCL1 ≥ 9.66 ng/ml in multivariable logistic regression analysis. Conclusion: SPARCL1 shows potential as novel biomarker of RV pathological remodelling and is associated with RV maladaptation and ventriculoarterial uncoupling in PH.

Alterations of right atrial (RA) function have emerged as determinants of outcome in pulmonary hypertension (PH). We aimed to clarify the pathophysiological associations of impaired RA conduit function with right ventricular (RV) function in PH. In 51 patients with PH (48 with pulmonary arterial hypertension), RA conduit function was assessed as echocardiographic peak early diastolic strain rate (PEDSR). PEDSR and cardiac magnetic resonance parameters were measured within 24 h of right heart catheterization and generation of pressure-volume loops to assess RV diastolic (RV end-diastolic pressure [EDP] and relaxation [Tau]) and systolic function. Spearman rho correlation and linear regression analysis were used to determine the association of PEDSR with RV function. The impact of PEDSR on time to clinical worsening was assessed using Kaplan-Meier and Cox regression analyses. Median (interquartile range) PEDSR was - 0.56 s -  1 (- 1.08 to - 0.37). Impaired PEDSR was significantly correlated with RV diastolic stiffness [EDP (rho = 0.570; p < 0.001) and Tau (rho = 0.500; p < 0.001)] but not with RV contractility or coupling. In multivariate linear regression including parameters of RV lusitropic and inotropic function, EDP remained independently associated with impaired PEDSR. During a median follow-up of 9 months, 23 patients deteriorated. After multivariate adjustment, PEDSR remained associated with clinical worsening (hazard ratio: 2.85; 95% confidence interval: 1.20-6.78). Altered RV lusitropy is associated with impaired RA conduit phase. PEDSR emerged as a promising, non-invasive, bedside-ready parameter to evaluate RV diastolic function and to predict prognosis in PH.

The functional relevance of right atrial (RA) function in pulmonary hypertension (PH) remains incompletely understood. The purpose of this study was to explore the correlation of cardiac magnetic resonance (CMR) feature tracking-derived RA phasic function with invasively measured pressure-volume (P-V) loop-derived right ventricular (RV) end-diastolic elastance ( E ed ) and RV-arterial coupling [ratio of end-systolic elastance to arterial elastance ( E es / E a )]. In 54 patients with severe PH, CMR was performed within 24 h of diagnostic right heart catheterization and P-V measurements. RA phasic function was assessed by CMR imaging of RA reservoir, passive, and active strain. The association of RA phasic function with indexes of RV function was evaluated by Spearman's rank correlation and linear regression analyses. Median [interquartile range] RA reservoir strain, passive strain, and active strain were 19.5% [11.0-24.5], 7.0% [4.0-12.0], and 13.0% [7.0-18.5], respectively. E es / E a was 0.73 [0.48-1.08], and E ed was 0.14 mmHg/mL [0.05-0.22]. RV diastolic impairment [RV end-diastolic pressure (EDP) and E ed ] was correlated with RA phasic function, but E a and E es were not. In addition, RA phasic function was correlated with inferior vena cava diameter. In multivariate linear regression analysis, adjusting for key P-V loop indexes, E ed and EDP remained significantly associated with RA phasic function. We conclude that RA phasic function is altered in relation to impaired diastolic function of the chronically overloaded right ventricle and contributes to backward venous flow and systemic congestion. These results call for more attention to RA function in the management of patients with PH. NEW & NOTEWORTHY There is growing awareness of the importance of the right atrial (RA)-right ventricular (RV) axis in pulmonary hypertension (PH). Our results uncover alterations in RA phasic function that are related to depressed RV lusitropic function and contribute to backward venous return and systemic congestion in chronic RV overload. Assessment of RA function should be part of the management and follow-up of patients with PH.