Pulmonary Circulation

Background: Pulmonary arterial hypertension (PAH) manifests by pulmonary vascular remodeling, caused by hyper-proliferation and apoptosis resistance of resident PA cells, elevated right ventricular (RV) afterload, and death of RV failure. The goal of this study is to determine the role and mechanisms of action of mitotic spindle-associated microtubule regulator DLG associated protein 5 (DLGAP5) in pulmonary vascular remodeling in PAH. Methods: DLGAP5 expression was assessed in human PAH transcriptomic datasets, PAH patients' lung tissues, pulmonary artery smooth muscle cells (PASMCs), and pulmonary artery adventitial fibroblasts (PAAFs). Loss-of-function, molecular, functional, RNAseq, and network analyses were performed. Smooth muscle (SM)-specific DLGAP5 knockdown using adeno-associated virus serotype 6 (AAV6) in mice with SU5416/hypoxia-induced PH was performed. Results: DLGAP5 was overaccumulated in small muscular PAs, PASMCs, and PAAFs from PAH lungs. siRNA DLGAP5 down-regulated cyclin A2, suppressed hyper-proliferation, increased pro-apoptotic BIM, and induced apoptosis in human PAH PASMCs and PAAFs. DEP domain containing 1 (DEPDC1) was identified and validated as a downstream effector of DLGAP5 in PAH PASMCs responsible for hyper-proliferation and apoptosis resistance of human PAH PASMCs. SM-specific DLGAP5 knockdown with AAV6-enTagln-shDLGAP5 reversed DEPDC1 overaccumulation in small muscular PAs, reduced PA medial thickness, RV systolic pressure, mean PA pressure, and RV hypertrophy in mice with SuHx-induced PH. Conclusions: DLGAP5 supports proliferation and survival of human PAH PASMCs and PAAFs through up-regulation of cyclin A2, DEPDC1, and suppression of BIM, and promotes PA remodeling and PH in mice. DLGAP5 could serve as a potential molecular target to attenuate pulmonary vascular remodeling in PAH.

Pulmonary arterial hypertension (PAH) is characterized by the hyperproliferative phenotype of pulmonary arterial smooth muscle cells (PAVSMCs) and extensive extracellular matrix (ECM) remodeling. Vacuolar protein sorting 34 (VPS34), a Class III Phosphatidylinositol-3 Kinase (PI3K), regulates cell proliferation, but its specific function in pulmonary vascular remodeling remains poorly understood. We aim to determine the role of VPS34 activation and regulation in PAVSMC proliferation, matrix deposition, and the progression of PAH. First, we found a marked decrease in the inhibitory phosphorylation of VPS34 at Ser164 (P-S164-VPS34), indicative of constitutive VPS34 activation, in remodeled pulmonary arteries (PAs) from human PAH patients and male mice exposed to Sugen/Hypoxia (SuHx). In human PAH PAVSMCs, siRNA-mediated knockdown of VPS34 or pharmacological inhibition via the selective inhibitor SAR405 significantly blunted cell proliferation and reduced the production of fibronectin, a major component of the remodeled ECM. Mechanistically, we found that Akt activation in PAH PAVSMCs promotes VPS34 activation; Protein Phosphatase 1 Catalytic Subunit Alpha is a potential effector in VPS34 activation; and active VPS34 led to the activation of the mTOR/S6 signaling pathway with tuberous sclerosis complex 2 downregulation. Finally, in vivo treatment with SAR405 attenuated right ventricular systolic pressure, right ventricular hypertrophy, and pulmonary vascular remodeling in SuHx mice, while concurrently decreasing vascular fibronectin deposition. Our study demonstrates that VPS34 activation, driven by an Akt-mediated loss of the inhibitory P-S164, is crucial in PAVSMC proliferation and fibrotic vascular remodeling in PAH. Targeting the VPS34 axis represents a promising anti-remodeling therapeutic strategy for the treatment of PAH. Keywords: Pulmonary arterial hypertension, vacuolar protein sorting 34, pulmonary vascular remodeling, pulmonary arterial vascular smooth muscle

BackgroundMacitentan is a dual endothelin receptor antagonist used in pulmonary arterial hypertension (PAH) and has also been studied in other forms of pulmonary hypertension (PH). Evidence across PH phenotypes has been heterogeneous. This study aims to evaluate the efficacy and safety of macitentan compared with concurrent control therapies across PH phenotypes and to explore whether treatment effects differ according to PH classification. MethodsPubMed, Embase, and the Cochrane database were searched from inception through June 2026. Randomized and non-randomized comparative studies of patients with PH receiving macitentan were eligible. Prespecified efficacy outcomes were 6-minute walk distance (6MWD), pulmonary vascular resistance (PVR), mean pulmonary arterial pressure (mPAP), and cardiac index; prespecified safety outcomes included mortality and adverse events. Random-effects meta-analyses were performed using mean differences (MDs) for continuous outcomes and risk ratios (RRs) for dichotomous outcomes. ResultsFourteen studies met the eligibility criteria. Macitentan was associated with a greater reduction in PVR (MD, -134.50 dyn.s.cm; 95% CI, -226.10 to -42.86) and an increase in cardiac index (MD, 0.44 L/min/m; 95% CI, 0.22 to 0.67). No statistically significant difference was observed for 6MWD (MD, 5.04 m; 95% CI, -15.06 to 25.14) or mPAP (MD, -2.70 mmHg; 95% CI, -6.46 to 1.05). In pooled safety analyses, mortality was lower and risks of anemia, peripheral edema, and discontinuation due to adverse events were higher with macitentan. Heterogeneity and the small number of studies within several phenotype-specific analyses limited the precision of subgroup findings. ConclusionsMacitentan-containing therapy was associated with favorable changes in selected hemodynamic outcomes but not with a statistically significant improvement in 6MWD or mPAP. The safety findings were characterized by higher risks of anemia, peripheral edema, and treatment discontinuation. These results should be interpreted in the context of differences in PH phenotype, dose, comparator, study design, and outcome measurement.

Rationale: Pulmonary hypertension is defined by pulmonary hemodynamics, but diagnostic and prognostic biomarkers remain limited. Nuclear magnetic resonance (NMR) spectroscopy provides detailed insights, particularly in the lipid metabolism. Objectives: To explore circulating NMR-derived metabolites and lipoprotein-related parameters for their association with pulmonary hemodynamics and to analyse their prognostic properties in pulmonary arterial hypertension (PAH). Methods: Retrospective analysis of a PAH cohort with complete diagnostic workup including right heart catheterization and baseline serum samples, from the prospective GRaz Pulmonary Hypertension-Metabolism (GRAPH-M) registry. Measurements: NMR-derived metabolites and lipoprotein-related parameters were analyzed for their association with clinically relevant parameters of PAH. We defined PHIHDL, a score derived from high-density lipoprotein (HDL) related measures based on their strong association with pulmonary hemodynamics, and evaluated its prognostic value. Results: We included 100 patients with PAH treated at the PH clinic of LKH University Hospital, Medical University of Graz, between 2011 and 2021. Age was 61{+/-}15 years, female/male ratio 2.5, BMI 26 {+/-}7 kg/m2, mPAP 41{+/-}16 mmHg, PAWP 8.8{+/-}3.2 mmHg, PVR 8.0{+/-}4.9 WU, and median survival was 8.0 years. During follow-up, 46 patients died. We identified a cluster of 12 HDL-related measures that showed significant inverse association to pulmonary hemodynamics and derived PHIHDL from the reversed scaled average of these particles. PHIHDL was associated with all-cause mortality after adjustment for age and sex (HR 2.96, 95% CI 1.52-5.70), independent of the clinical risk scores COMPERA 2.0 and REVEAL Lite2. Conclusion: PHIHDL, a pulmonary hemodynamics-based metabolomic score, provides independent prognostic information beyond established risk scores in PAH.

ABSTRACT Background: The modified Blalock-Taussig-Thomas shunt (mBTTS) sustains pulmonary blood flow in infants with cyanotic or single-ventricle heart disease, but shunt thrombosis occurs in 8-12% of cases and carries substantial mortality. Systemic anticoagulation has limited efficacy. Objectives: This study tested whether surgically modifiable shunt geometry might form a second, patient-specific approach to maintaining shunt patency, via management of thrombogenic hemodynamics. Methods: Three-dimensional mBTTS anatomies were reconstructed from computed tomography in 10 infants (4 thrombosed, 6 patent). Pulsatile inlet waveforms derived from Doppler ultrasound were coupled with three-element Windkessel outlet models individually calibrated to catheter-derived pressures. Transient computational fluid dynamics simulations predicted wall shear rate (WSR) and elongational strain rate (ESR). Associations between surgically relevant geometric parameters and hemodynamic metrics were evaluated. Results: Simulated pressures closely matched clinical measurements at all four outlets in all 10 models. Despite substantial anatomic variability, peak WSR and ESR consistently localized to the shunt-subclavian junction in 8 of 10 patients, including all 4 thrombosed shunts. Greater deviation of shunt insertion angle from perpendicular was associated with higher systolic and cycle-averaged normalized WSR ({rho} = 0.94, p < 0.001; {rho} = 0.88, p = 0.002) and ESR ({rho} = 0.82, p = 0.007; {rho} = 0.70, p = 0.03). Larger shunt diameter was associated with lower WSR and ESR. Thrombosed shunts demonstrated higher shear-related metrics than patent shunts, particularly during systole. Conclusions: In patient-specific mBTTS anatomies, insertion angle and shunt diameter are determinants of local abnormal flow, concentrated at the shunt-subclavian junction. Patient-specific hemodynamic assessment may inform shunt construction and interstage risk stratification.

Background: Pulmonary vascular disease identifies an important phenotype of heart failure with preserved ejection fraction (HFpEF), but the relationship between systemic leukocyte activation and invasively measured pulmonary vascular hemodynamics is incompletely understood. We investigated whether the neutrophil-to-lymphocyte ratio (NLR) is associated with combined post- and precapillary pulmonary hypertension (CpcPH), pulmonary vascular burden, and subsequent cardiovascular events in HFpEF. Methods: We retrospectively studied 169 adults with HFpEF and invasively confirmed postcapillary pulmonary hypertension undergoing clinically indicated right heart catheterization. CpcPH was defined as pulmonary vascular resistance (PVR) >2 Wood units. Associations between NLR and CpcPH were evaluated using multivariable logistic regression. Fine-Gray competing-risk models assessed the composite of cardiovascular death or first heart failure hospitalization. Results: Overall, 80 patients had isolated postcapillary pulmonary hypertension and 89 had CpcPH. PVR increased across ascending NLR tertiles (1.8 [1.4-2.5], 2.2 [1.6-3.1], and 2.6 [1.7-3.8] Wood units; P for trend=0.002), whereas pulmonary artery wedge pressure showed no significant trend. Compared with the lowest NLR tertile, the highest tertile was associated with greater odds of CpcPH after adjustment for clinical covariates and filling pressure (adjusted odds ratio, 2.47 [95% CI, 1.01-6.01]; P=0.047). Higher NLR tertiles were also associated with a graded increase in cardiovascular events (Gray's P<0.001). The highest NLR tertile remained associated with outcome after adjustment for clinical and hemodynamic variables (adjusted subdistribution hazard ratio [sHR], 2.37 [95% CI, 1.12-5.03]; P=0.024). Conclusions: Among patients with HFpEF and postcapillary pulmonary hypertension, higher NLR was associated with greater pulmonary vascular hemodynamic burden, CpcPH, and subsequent cardiovascular risk. NLR may provide a readily available marker of a high-risk HFpEF phenotype warranting prospective validation.

Pulmonary hypertension (PH) is a progressive and fatal disease characterized by pulmonary vascular remodeling, inflammation, and immune dysregulation. Myeloid-derived suppressor cells (MDSCs) and macrophages contribute to PAH pathobiology; however, the molecular regulators of their pathological activation remain poorly defined. The triggering receptor expressed on myeloid cells 2 (TREM2) is an immunomodulatory receptor that shapes myeloid cell metabolism, survival, and immunosuppressive function, yet its role in PH has not been investigated. Methods: Single-cell RNA sequencing (scRNA-seq) data from human pulmonary artery tissue (GSE210248; n=3 PAH, n=3 donors) were analyzed to characterize TREM2 expression across cell populations. Wild-type (WT) and global TREM2 knockout (TREM2 KO) mice were exposed to chronic hypoxia (10% FiO2, 28 days). Hemodynamic, histological, flow cytometric, and ex vivo functional assessments were performed. TREM2 expression was measured by flow cytometry in circulating MDSCs from PAH patients (n=22) and healthy controls (n=13). Results: TREM2 was markedly enriched in monocyte/macrophage populations in PH pulmonary arteries and TREM2-high immune cells exhibited transcriptional downregulation of chemotaxis programs and upregulation of antigen processing and MHC II presentation pathways. TREM2 deficiency significantly attenuated hypoxia-induced RVSP increase, right ventricular dysfunction, pulmonary inflammation, and vascular remodeling. MDSCs from TREM2 KO mice exerted significantly less suppression of CD4+ and CD8+ T cell proliferation. TREM2 was significantly elevated in circulating MDSCs from PAH patients and showed a directional association with hemodynamic severity. Conclusions: TREM2 is a novel regulator of myeloid-driven immunosuppression and vascular remodeling in PAH and warrants investigation as a therapeutic target and biomarker.

BackgroundPulmonary hypertension (PH) involves progressive vascular remodeling and perivascular inflammation. Despite modest clinical improvements with current therapies, their limited ability to reverse remodeling or restore immune homeostasis highlights the need for novel agents. Liriodendrin (Lidd), derived from Sargentodoxae caulis, exhibits anti-inflammatory and antiproliferative activities, but its efficacy and molecular targets in PH are unknown. MethodsTwo well-established PH animal models - the SU5416/hypoxia (SuHx) mice model and monocrotaline (MCT)-induced rat model - were employed for in vivo assessment of Lidd conducted pharmacological effects. Primary human pulmonary artery smooth muscle cells (hPASMCs) were utilized for mechanistic investigations. RNA-sequencing (RNA-seq) analysis was conducted to identify potential signaling pathways modulated by Lidd treatment. The direct molecular target of Lidd was determined through integrated application of drug affinity responsive target stability (DARTS) assay coupled with western blot validation. To delineate histone lactylation-mediated transcriptional regulation, we combined Cleavage Under Targets and Tagmentation (CUT&Tag) sequencing data analysis followed by chromatin immunoprecipitation quantitative PCR (ChIP-qPCR) verification. Genetic validation was achieved using PFKFB3-deficient murine models to verify the specificity of Lidd-mediated pharmacological actions. ResultsLidd administration attenuated pulmonary vascular remodeling, perivascular macrophage infiltration and PH progression in both SuHx and MCT models. Transcriptomic profiling of Lidd-treated hPASMCs revealed predominant enrichment of downregulated genes in inflammatory and cytokine-associated pathways. Mechanistically, Lidd directly bound PFKFB3 and enhanced its interaction with FZR1, promoting PFKFB3 ubiquitination and degradation, which reduced glycolysis-driven lactate and consequent histone lactylation. This, in turn, diminished transcriptional activation of proliferative and inflammatory mediators, including CCND1, TNC, and CCL2. Notably, exogenous lactate supplementation or endogenous lactate accumulation restored histone lactylation and paradoxically potentiated Lidds inhibitory effects on PASMC proliferation and migration, whereas p300 inhibition abrogated these lactate-mediated effects. Importantly, Lidd failed to confer additional protection in PFKFB3-deficient mice, confirming PFKFB3 as the primary target mediating its therapeutic action. ConclusionOur findings reveal that Lidd selectively targets the PFKFB3-mediated glycolytic-epigenetic axis to suppress PASMC phenotypic transformation and pulmonary vascular remodeling, positioning it as a promising therapeutic candidate for PH.

Pericytes are essential regulators of pulmonary vascular homeostasis, but their isolation from rat lung tissue is challenging because no single marker uniquely identifies them and contaminating fibroblasts, endothelial cells, hematopoietic cells, and vascular smooth muscle cells can persist during isolation and culture. Here, we describe a rapid and efficient protocol for the isolation of primary rat pulmonary pericytes using sequential magnetic depletion of CD45-positive hematopoietic cells and CD31-positive endothelial cells, followed by positive selection for NG2-positive cells. The isolated cells were expanded in culture and characterized by immunofluorescence and functional co-culture assays. Cultured cells displayed typical pericyte morphology and expressed the pericyte-associated markers NG2, PDGFR{beta}, and 3G5, with minimal expression of CD31, CD45, PDGFR and MYH11. In endothelial cell-pericyte ECM gel co-culture assays, isolated pericytes associated with endothelial cords and were frequently observed near network branch points and junctions, further supporting their pericyte identity. This method yields an enriched population of primary pulmonary pericytes suitable for downstream applications, including cell culture and functional studies. Overall, this streamlined protocol provides a practical platform for studying pulmonary pericyte biology in rat models of health and vascular disease.

Background: Heritable thoracic aortic aneurysms and dissections (HTAD) are caused by rare variants in up to 42 genes, but many affected individuals and families remain genetically unsolved. We sought to determine if aortic phenotypes occurred in cases with de novo pathogenic variants in known syndromic genes. Methods: We queried a clinical exome/genome sequencing database for individuals with de novo pathogenic or likely pathogenic variants (PVs) and aortic Human Phenotype Ontology terms. Genes with PVs present in three or more unrelated cases with aortic phenotypes were further evaluated using published evidence, genome-wide rare-variant burden analyses in the UK Biobank and All of Us, and case-level data from independent thoracic aortic disease (TAD) cohorts. Results: Among 250 individuals with de novo PVs and aortic phenotypes, 32 (12.8%) had PVs in genes included on commercial HTAD panels and were excluded from further analyses. The remaining 218 individuals had PVs in 144 genes, including 16 genes identified in three or more unrelated individuals. Genes involved in chromatin regulation and RAS/mitogen-activated protein kinase (MAPK) signaling, as well as several genes associated with overgrowth syndromes, were recurrently observed. Across biobank analyses, independent TAD cohorts, and published reports, the strongest evidence for aortic involvement was observed for DNMT3A, NF1, PTPN11, PPP1CB, NSD1, ANKRD11, ADNP, and ABL1, including significant rare-variant burden associations for three of these genes. Aortic phenotypes also occurred across multiple Coffin-Siris/BAF-complex genes. Conclusions: Aortic phenotypes occur across a diverse range of genetic syndromes beyond established HTAD genes. These findings support broader genomic testing in TAD, expansion of HTAD panels to include additional syndromic genes, and consideration of aortic surveillance in select syndromic disorders.

Objective(s)Tricuspid valve (TV) remodeling and functional tricuspid regurgitation (FTR) progression during right ventricular (RV) pressure overload and reverse remodeling after resolution of RV afterload is poorly understood. We set out to investigate tricuspid leaflet tissue response to induction and subsequent alleviation of pressure overload in a large animal model of RV failure with FTR. MethodsFifteen healthy adult male Dorset sheep (72{+/-}4 kg) underwent pulmonary artery banding (PAB) to induce RV failure and FTR. After 8 weeks, 7 sheep (PAB, n=7) were terminated, and remaining 8 had the PAB removed (rPAB, n=8) and were followed for another 8 weeks before termination. Both groups underwent epicardial echocardiography and hemodynamic assessment during banding surgery and at terminal operation. Ten healthy sheep served as a control group (CTL, n=10) and underwent terminal procedure only. In all animals, TV leaflets and right ventricular (RV) tissue were harvested at terminal procedure and analyzed histologically and transcriptionally. ResultsTV leaflets in PAB animals showed increased cross-sectional area and ECM alterations, some of which persisted after resolution of RV pressure overload. rPAB valves exhibited distinct ECM composition, with notably altered mucin and fibrin content, suggesting a shift toward matrix stabilization, dissimilar to control and PAB. RNA sequencing uncovered a unique molecular state in rPAB valves, with persistent changes in PRG4, PDE3A, CXCL8, and HLA transcripts. RV tissue also demonstrated a separate remodeling trajectory, with sustained expression of stress-related genes including PDE3A, NAV2, ANFB, and ACTS. These findings indicate that both valve and ventricular tissues retain a persistent remodeled phenotype post-unloading. ConclusionsTV leaflets actively remodel in response to hemodynamic stress and do not fully revert to a normal state after relief of pressure overload. This persistent altered phenotype may represent a biological contribution of the TV leaflets to recurrent TR with implications for long-term outcomes following treatment of FTR. Clinical Perspective What is new?O_LIRelief of right ventricular pressure overload, in a large animal model, resulted in substantial reverse remodeling of the right heart and reduction of tricuspid regurgitation severity, but tricuspid valve leaflets did not return to a normal state. C_LIO_LIReverse remodeled leaflets remained enlarged despite normalization of hemodynamics with an altered extracellular matrix. C_LIO_LICellular proliferation and immune cell infiltration observed during pressure overload resolved after unloading, yet transcriptomic analysis identified a distinct molecular phenotype that differed from both healthy and diseased valves. C_LIO_LITricuspid valve leaflets are active biological participants in the remodeling process and exhibit persistent adaptation or maladaptation after resolution of the initiating hemodynamic stress. C_LI What Are the Clinical Implications?O_LISecondary tricuspid regurgitation should be considered a disease involving both right heart geometry and leaflet biology. C_LIO_LIResolution of the underlying cause of tricuspid regurgitation may not restore leaflet structure and molecular homeostasis. C_LIO_LIPersistent leaflet remodeling may contribute to residual or recurrent tricuspid regurgitation despite successful treatment of pulmonary hypertension or other inciting conditions. C_LIO_LITherapies directed at leaflet remodeling may ultimately complement surgical and transcatheter strategies currently focused on annular and ventricular geometry. C_LI

Background: Cyclooxygenase (COX) inhibitors have been used to treat hemodynamically significant patent ductus arteriosus (PDA) in preterm infants. If the PDA remains hemodynamically significant after an initial course, additional courses of COX inhibitors or surgical intervention are considered. This systematic review evaluates PDA closure rates and safety after repeated courses of COX inhibitors in preterm infants. Methods: Embase, Medline, and two other databases were searched through April 2026 (pre-registered in PROSPERO [CRD42023454003]). Studies reporting PDA closure rates after two or more courses of COX inhibitors were included. Risk of bias was assessed using Joanna Briggs Institute tools. Meta-analyses with random-effects models were performed to calculate the proportions of PDA closure. Multilevel mixed-effects meta-regression was used to compare closure rates across treatment courses while accounting for within-study correlations. Results: A total of 33 studies were included. The PDA closure rates after the first, second, and third courses of COX inhibitors were 62% (95% confidence interval [CI] 57-67%), 46% (95% CI 40-52%; p < 0.001 vs the first course), and 38% (95% CI 27-51%; p = 0.001 vs the first course), respectively. Lower closure rates with repeated courses were also observed across most subgroups (extremely preterm/extremely low birth weight, ibuprofen, indomethacin, and intravenous administration), except for the second course of oral administration. Data on adverse events were limited, preventing meta-analysis. Conclusions: PDA closure rates after the second and third courses of COX inhibitors were lower than those after the first course in preterm infants. Evidence regarding the safety of repeated courses remains limited.

BackgroundRight ventricular (RV) adaptation determines outcomes in pulmonary arterial hypertension (PAH), yet multicellular molecular programs associated with adaptive versus maladaptive RV remodeling in living humans remain incompletely defined. MethodsWe collected 32 human RV tissue biopsies from patients with idiopathic PAH, systemic sclerosis-associated PAH (SSc-PAH), systemic sclerosis without pulmonary hypertension, with 24 nonfailing donor RVs serving as controls. We performed single-nucleus RNA sequencing and integrated cell-type specific transcriptional programs with contemporaneously obtained multi-beat pressure-volume loop measurements of RV contractility (Ees, end-systolic elastance) and RV-pulmonary arterial coupling (the ratio of Ees to Ea, the effective arterial load). SSc modification of PAH-associated biology was assessed using interaction terms. Bulk RV proteomic pathway enrichment was performed to assess an orthogonal molecular layer, and exploratory cell-cell communication analyses alongside independent spatial transcriptomic analyses were performed to contextualize key findings. ResultsPAH was associated with broad depletion of biosynthetic, trafficking, and mitochondrial programs across cell types. SSc modified the magnitude of many PAH-associated transcriptional programs while largely preserving pathway directionality. Significant multicellular pathway enrichments were associated with RV-pulmonary arterial coupling. Joint analysis of Ees, Ea, and Ees/Ea identified biologic programs associated with different RV responses to varying loading conditions. Preserved coupling was characterized by enriched extracellular matrix, laminin-integrin, receptor tyrosine kinase, mitochondrial, and translational programs involving fibroblast, endothelial, endocardial, and cardiomyocyte compartments. Cell- cell communication analyses predicted coordinated stromal-vascular signaling networks involving laminin-integrin and endothelial-to-mural signaling in preserved coupling. Proteomic and spatial analyses supported recurrent multicellular themes. ConclusionsRV adaptation in PAH is associated with distinct, coordinated multicellular programs that vary with load and contractile response. RV-PA coupling in PAH is associated with multicellular remodeling that extends beyond cardiomyocytes and reflects organized vascular-stromal support architecture. These findings identify extracellular matrix, laminin- integrin signaling, mitochondrial, and translational programs as associated with adaptive RV remodeling in PAH. Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABSO_LICell type-resolved molecular profiling of living human RV tissue identifies PAH-associated depletion of biosynthetic, trafficking, mitochondrial, and repair-associated programs across multiple cardiac cell types. C_LIO_LIIntegration with contemporaneously obtained pressure-volume loop physiology demonstrates that preserved RV-pulmonary arterial coupling under lower load was associated predominantly with cardiomyocyte mitochondrial and metabolic competency, whereas preserved coupling under higher load was associated with extracellular matrix remodeling and vascular-stromal signaling. C_LIO_LIProteomic, cell-cell communication, and spatial analyses provided orthogonal support for coordinated extracellular matrix and vascular-stromal programs associated with preserved RV-pulmonary arterial coupling. C_LI What are the clinical implications?O_LIThese findings shift the biology of RV adaptation from a predominantly cardiomyocyte- centered model toward a multicellular tissue model in which metabolic, matrix, and vascular support programs vary according to loading conditions and contractile states. C_LIO_LIExtracellular matrix-integrin signaling, endothelial-mural communication, and mitochondrial competency represent candidate pathways for mechanistic investigation toward RV-directed therapies in PAH. C_LI

Objective: Lymphatic dysfunction has been implicated in exacerbating fibrosis in numerous diseases, yet the role of the lymphatic system in laryngotracheal injury has not previously been explored. This study aims to evaluate lymphatic vascular remodeling in a murine model of laryngotracheal stenosis (LTS) and determine how pharmacologic blockade of lymphangiogenesis impacts airway healing after mucosal injury. Methods: LTS was induced in C57BL6 mice using an established chemomechanical injury model. Lymphatic density was quantified using LYVE-1 immunohistochemistry. Mice were treated with the VEGFR-3-selective tyrosine kinase inhibitor SAR131675 to block lymphangiogenesis after injury. Outcomes assessed included survival, histopathology, immunohistochemistry, and Evans blue dye vascular leakage. Results: Laryngotracheal injury induced a substantial increase in subepithelial lymphatic vessel density concomitant with fibrotic remodeling. Pharmacologic inhibition of VEGFR-3 signaling with SAR131675 abrogated this lymphangiogenic response and resulted in markedly increased mortality, impaired epithelial repair with obstructive sloughing, increased edema, and persistent histopathologic evidence of tissue injury. A qualitative increase in pathologic fibrocellular remodeling was also observed, though with no measurable difference in lamina propria thickness. Conclusion: These findings establish lymphatic remodeling as an essential component of successful airway repair following mucosal injury. Lymphatic dysfunction is a common feature of known risk factors for LTS including diabetes, obesity, and prematurity, and can be exacerbated by positive pressure ventilation. Disruption of the lymphangiogenic response to airway injury may lead to stasis of pro-inflammatory factors that result in chronic inflammation, maladaptive remodeling, and pathologic tissue changes. The lymphatic vasculature is a viable target for future mechanistic study and potential therapeutic intervention following airway injury.

Genetic testing for cardiomyopathy and arrhythmia (CM/ARRH) provides diagnostic information and informs screening for at-risk relatives. Clinical guidelines recommend genetic testing for these conditions; however, data on how genetic test results influence clinical management recommendations are limited. Here, we determined the frequency of cardiologist-recommended management changes for patients following CM/ARRH genetic testing. This was a retrospective cross-sectional study of patients referred for multigene panel testing between April 2016 and April 2024. Genetically-experienced cardiologists at multicenter academic clinical practices were recruited for participation to complete surveys indicating clinical decision making on patients who had genetic testing. Cases for review were randomly selected to have both positive and non-positive results. Among 249 patients (138 positive, 111 non-positive), 136 (54.6%) received clinical management recommendations for their own or their at-risk relatives? care. Of these, 75 (55.1%) received recommendations for the patient?s own care, most frequently additional diagnostic tests/procedures (n=33). Compared to non-positive results, patients with positive results were more likely to receive recommendations for their own management (66/138, 47.8% vs 9/111, 8.1%; P<0.00001). Patients with positive results in arrhythmogenic cardiomyopathy genes had 263% higher odds of recommended management changes compared to those with TTN (OR=3.63, CI:1.40-9.84, P=0.009). The results from CM/ARRH genetic testing on affected patients influenced cardiologists? medical decision making and management recommendations. Additional research is needed to evaluate how genetic testing for CM/ARRH impact health outcomes.

BackgroundPulmonary hypertension (PH) is characterized by pulmonary vascular remodeling and smooth muscle cell accumulation, but the progenitor-like cells that contribute to this process remain incompletely defined. MethodsWe combined analyses of human pulmonary arterial hypertension lungs and experimental PH models with bulk and single-cell RNA sequencing, lineage tracing, inducible ablation of CD133+ cells, and conditional deletion of Cxcr4 in CD133+ cells. ResultsCD133 expression was markedly increased in human and experimental PH lungs. Transcriptomic analyses identified inflammatory, metabolic, chemokine-associated, and smooth muscle cell-like programs in CD133+ cells from PH lungs. Lineage tracing showed that CD133+ cells contributed to endothelial and smooth muscle cell compartments during experimental PH. Genetic ablation of CD133+ cells attenuated hypoxia-induced PH and pulmonary vascular remodeling, whereas Cxcr4 deletion in CD133+ cells reduced PH severity. ConclusionsCD133+ progenitor cells are functional contributors to pulmonary vascular remodeling, and CXCR4 signaling mediates their pathogenic activity. Targeting pathogenic CD133+ cell states or CXCL12/CXCR4 signaling may provide a strategy to limit vascular remodeling in PH.

Background: Adults with congenital heart disease (ACHD) represent a rapidly expanding population with evolving mortality patterns. Despite improved survival, excess mortality persists. Objective: To evaluate the incidence, causes, and predictors of mortality in a contemporary ACHD cohort. Methods: We performed a retrospective cohort study of adults (?18 years) first evaluated at Mayo Clinic from 2002?2023. Baseline clinical, imaging, and electrocardiographic data were analyzed. Vital status was determined using institutional records and the Accurint national mortality database. Kaplan-Meier analysis and Cox proportional hazard models were used to evaluate mortality and identify independent predictors of mortality Results: A total of 7,678 ACHD patients were included, with median age of 36.8 years and median follow-up of 11.4 years. During 78,768 patient-years of follow-up, 1,116 patients died (median age at death 57.2 years), corresponding to an annual mortality rate of 1.4%. The cumulative rate of all-cause mortality at 5, 10, 15, and 20 years was 6.9%, 12.0%, 19.0%, and 26.2%, respectively. When stratified by CHD complexity, the annual death rate in patients with severe CHD (2.4%/year) was twice that of patients with moderate or mild CHD (both 1.2%/year). Older age and ACHD subtypes, specifically, cyanotic heart disease (HR 3.9, 95% CI 2.9?5.3) and Fontan physiology (HR 3.2, 95% CI 2.3?4.4), were strongly associated with increased mortality. Additional independent predictors included male sex, ventricular dysfunction, advanced NYHA class, prior heart failure hospitalization, hypertension, smoking, coronary artery disease, renal dysfunction, and abnormal hemoglobin levels. Cardiovascular causes accounted for 57.7% of deaths with known etiology, predominantly heart failure (48.9%) and sudden cardiac death (21.9%), while non-cardiovascular causes were driven mainly by infection and malignancy. Conclusions: In this large contemporary ACHD cohort, mortality was driven by ventricular dysfunction, heart failure, and systemic end-organ involvement in addition to the underlying congenital anatomy. Both cardiovascular and non-cardiovascular causes contributed significantly to mortality. These findings underscore the need for comprehensive multidisciplinary ACHD care focused on early recognition of cardiac functional decline, management of acquired comorbidities, and end-organ dysfunction.

Background Mitral valve prolapse (MVP) is a leading cause of mitral regurgitation in high-income countries. Although the course of MVP is often benign, a subset of patients progresses to significant valvular insufficiency, and a smaller subset develops ventricular arrhythmias or sudden cardiac death. This study aimed to identify genetic variants associated with MVP and/or mitral annulus disjunction (MAD) using whole genome sequencing (WGS) in a Norwegian cohort. Methods WGS was performed in 93 individuals diagnosed with MVP and/or MAD, of whom 92 passed genomic quality control criteria. Variants were called using Genome Analysis ToolKit (GATK) Best Practices and compared with non Finnish European reference data from gnomAD (n = 34 029). Analyses were restricted to curated gene panels comprising: (i) non syndromic MVP and cardiomyopathy genes, (ii) syndromic/connective tissue disorder genes, and (iii) arrhythmia related genes. Enriched or underrepresented variants were identified using Fisher's exact test with Bonferroni correction for multiple testing. Independent variants were derived through linkage disequilibrium pruning. A sensitivity analysis was conducted using low coverage WGS data from an independent large Norwegian cohort -- the Trondelag Health Study (HUNT). Results Across predefined gene panels, several coding and regulatory variants were significantly associated with MVP and/or MAD. A missense variant in FLNA and regulatory variants in TBX5 and SMAD4 were enriched in these patients. Additional significant associations involved regulatory variants in XYLT1, HS3ST4, SH3PXD2B, and a synonymous variant in COL1A2. Furthermore, several regulatory variants in arrhythmia related genes (KCNK3, CACNA1D, NR2F1, KCNJ4 and others) were identified. Sensitivity analyses revealed population level differences, underscoring the need of cautious interpretation of these findings. Conclusion This targeted WGS study expands the genetic landscape of MVP and/or MAD, identifying several coding and regulatory variants across extracellular matrix, developmental, and electrophysiological pathways. These results support a multifactorial genetic architecture of MVP.

Background Exercise intolerance is common in adults with repaired tetralogy of Fallot (rTOF) despite preserved left ventricular ejection fraction (LVEF [&ge;]50%). Whether reduced exercise capacity is associated with early cardiac remodeling remains unclear. Objectives To determine whether reduced exercise capacity in rTOF with preserved LVEF is associated with diastolic dysfunction, atrial remodeling, right ventricular (RV) dysfunction, and myocardial fibrosis. Methods We retrospectively studied adults with rTOF and preserved LVEF who underwent cardiopulmonary exercise testing (CPET) and transthoracic echocardiography (TTE) and/or cardiac MRI (CMR) within 18 months. Exercise capacity was assessed by percent-predicted peak VO2 (ppVO2). Diastolic function and atrial remodeling were evaluated by TTE, and CMR assessed RV function and myocardial fibrosis. Results Reduced exercise capacity was associated with larger left atrial volume index (LAVI; p < 0.001), elevated E/e' and reduced e' velocity (both p < 0.05), and reduced RV systolic function (p < 0.001). LAVI correlated inversely with ppVO2 ({rho} = -0.27, p = 0.003). A composite diastolic dysfunction score showed a graded relationship with exercise capacity, with patients exhibiting [&ge;]2 abnormalities having lower ppVO2 than those with [&le;]1 abnormality (both p < 0.01). In contrast, pulmonary regurgitation (PR) severity and myocardial fibrosis by late gadolinium enhancement (LGE) were not associated with exercise capacity. Conclusions In adults with rTOF and preserved LVEF, reduced exercise capacity is associated with atrial remodeling, diastolic dysfunction, and RV dysfunction despite the absence of overt myocardial fibrosis. This suggests that multimodal imaging identifies an imaging-defined cardiac remodeling phenotype associated with early functional impairment.

Introduction: The key role of maladaptive renal sympathetic activation in hypertension has led to the development of catheter-based renal denervation therapies. Unfortunately, the lack of practical physiological biomarkers impairs patient selection and prevents intraprocedural feedback for renal denervation. Methods: We tested whether renal arterial hemodynamic measures and the beat-to-beat variability of these measures reflect renal sympathetic innervation in two tightly controlled preclinical models of renal denervation. Bilateral renal hemodynamic data were obtained from ten conscious unilaterally denervated rabbits and ten anesthetized unilaterally denervated pigs that subsequently underwent stepwise catheter-based radiofrequency denervation of the contralateral kidney. Renal arterial mechanics were assessed by quantification of wave speed and input impedance modulus and phase shift. Variability was quantified as the within-recording standard deviation of each metric when measured on a beat-to-beat basis. Results: Wave speed trended down after surgical denervation in rabbits (P = 0.054) but not swine (P = 0.57); wave speed variability was unchanged in both models. Renal arterial input impedance modulus was not significantly affected by surgical denervation. Surgical renal denervation increased input impedance phase shift in rabbits (P = 0.0048) but not in swine (P = 0.85). The beat-to-beat variability of modulus did not differ significantly between innervated and surgically denervated kidneys in either species. Surgical renal denervation decreased phase shift variability in swine (P = 0.0048) but not rabbits. The difference in phase shift variability between kidneys in swine was eliminated after a single round of branch-vessel ablation (P = 0.42). Discussion: Renal arterial wave speed, input impedance, and their beat-to-beat variability did not consistently or dose-dependently reflect renal sympathetic innervation in these models. These findings argue against their use as direct physiological surrogates for renal sympathetic outflow.