Promising dawn in the management of pulmonary hypertension: The mystery veil of gut microbiota.

The gut microbiota is a complex community of microorganisms inhabiting the intestinal tract, which plays a vital role in human health. It is intricately involved in the metabolism, and it also affects diverse physiological processes. The gut-lung axis is a bidirectional pathway between the gastrointestinal tract and the lungs. Recent research has shown that the gut microbiome plays a crucial role in immune response regulation in the lungs and the development of lung diseases. In this review, we present the interrelated factors concerning gut microbiota and the associated metabolites in pulmonary hypertension (PH), a lethal disease characterized by elevated pulmonary vascular pressure and resistance. Our research team explored the role of gut-microbiota-derived metabolites in cardiovascular diseases and established the correlation between metabolites such as putrescine, succinate, trimethylamine N-oxide (TMAO), and N, N, N-trimethyl-5-aminovaleric acid with the diseases. Furthermore, we found that specific metabolites, such as TMAO and betaine, have significant clinical value in PH, suggesting their potential as biomarkers in disease management. In detailing the interplay between the gut microbiota, their metabolites, and PH, we underscored the potential therapeutic approaches modulating this microbiota. Ultimately, we endeavor to alleviate the substantial socioeconomic burden associated with this disease. This review presents a unique exploratory analysis of the link between gut microbiota and PH, intending to propel further investigations in the gut-lung axis.

Isolated Perfused Hearts for Cardiovascular Research: An Old Dog with New Tricks.

In modern cardiovascular research, isolated perfused hearts have become cost-effective and highly reproducible tools to investigate the mechanisms of cardiovascular diseases (CVDs). Since they were first introduced in the nineteenth century, isolated perfused hearts have been extensively used for testing novel therapies, elucidating cardiac metabolic and electrophysiological activities, and modeling CVDs, including ischemic heart disease, arrhythmias, and hyperacute rejection. In recent years, ex vivo heart perfusion (EVHP) has shown potential in cardiac transplantation by allowing prolonged preservation and reconditioning of donor hearts. In this review, we summarize the evolution of the isolated perfused heart technique and its applications in cardiovascular research to help researchers comprehensively understand the capabilities of isolated heart models and provide guidance to use them to investigate various CVDs.

Development and Validation of a Novel Predictive Model for the Early Differentiation of Cardiac and Non-Cardiac Syncope.

Background: The diagnosis of cardiac syncope remains a challenge. This study sought to develop and validate a diagnostic model for the early identification of individuals likely to have a cardiac cause. Methods: 877 syncope patients with a determined cause were retrospectively enrolled at a tertiary heart center. They were randomly divided into the training set and validation set at a 7:3 ratio. We analyzed the demographic information, medical history, laboratory tests, electrocardiogram, and echocardiogram by the least absolute shrinkage and selection operator (LASSO) regression for selection of key features. Then a multivariable logistic regression analysis was performed to identify independent predictors and construct a diagnostic model. The receiver operating characteristic curves, area under the curve (AUC), calibration curves, and decision curve analysis were used to evaluate the predictive accuracy and clinical value of this nomogram. Results: Five independent predictors for cardiac syncope were selected: BMI (OR 1.088; 95% CI 1.022-1.158; P =0.008), chest symptoms preceding syncope (OR 5.251; 95% CI 3.326-8.288; P <0.001), logarithmic NT-proBNP (OR 1.463; 95% CI 1.240-1.727; P <0.001), left ventricular ejection fraction (OR 0.940; 95% CI 0.908-0.973; P <0.001), and abnormal electrocardiogram (OR 6.171; 95% CI 3.966-9.600; P <0.001). Subsequently, a nomogram based on a multivariate logistic regression model was developed and validated, yielding AUC of 0.873 (95% CI 0.845-0.902) and 0.856 (95% CI 0.809-0.903), respectively. The calibration curves showcased the nomogram's reasonable calibration, and the decision curve analysis demonstrated good clinical utility. Conclusion: A diagnostic tool providing individualized probability predictions for cardiac syncope was developed and validated, which may potentially serve as an effective tool to facilitate early identification of such patients.

Circulating acetylcholine serves as a potential biomarker role in pulmonary hypertension.

BACKGROUND: An increased acetylcholine (ACh) level in the right ventricle tissue of pulmonary hypertension (PH) was revealed, which indicated the important role of ACh in disease pathogenesis. However, the relationship between plasma ACh levels and disease conditions and patients' prognosis has not been investigated. We aimed to explore the association between plasma ACh levels and the prognosis of patients with PH. We also discussed the feasibility of plasma ACh as a biomarker, which may contribute to the management of PH patients in the future. METHODS: Patients with confirmed PH in Fuwai Hospital from April 2019 to August 2020 were enrolled. The primary clinical outcome in this study was defined as a composite outcome, including death/lung transplantation, heart failure, and worsening of symptoms. Fasting plasma was collected to detect the ACh levels. The association between ACh levels and patients' prognosis was explored. RESULTS: Finally, four hundred and eight patients with PH were enrolled and followed for a mean period of 2.5 years. Patients in the high ACh group had worse World Health Organization Functional Class (WHO-FC), lower 6-minute walk distance (6 MWD), and higher N-terminal pro-brain natriuretic peptide (NT-proBNP). Notably, echocardiographic and hemodynamic parameters in the high metabolite group also suggested a worse disease condition compared with the low ACh group. After adjusting for confounders, compared with low ACh patients, those with high metabolite levels still have worse prognoses characterized as elevated risk of mortality, heart failure, and symptoms worsening. CONCLUSION: High circulating ACh levels were associated with severe PH conditions and poor prognosis, which might serve as a potential biomarker in PH.

Single-cell RNA sequencing in donor and end-stage heart failure patients identifies NLRP3 as a therapeutic target for arrhythmogenic right ventricular cardiomyopathy.

BACKGROUND: Dilation may be the first right ventricular change and accelerates the progression of threatening ventricular tachyarrhythmias and heart failure for patients with arrhythmogenic right ventricular cardiomyopathy (ARVC), but the treatment for right ventricular dilation remains limited. METHODS: Single-cell RNA sequencing (scRNA-seq) of blood and biventricular myocardium from 8 study participants was performed, including 6 end-stage heart failure patients with ARVC and 2 normal controls. ScRNA-seq data was then deeply analyzed, including cluster annotation, cellular proportion calculation, and characterization of cellular developmental trajectories and interactions. An integrative analysis of our single-cell data and published genome-wide association study-based data provided insights into the cell-specific contributions to the cardiac arrhythmia phenotype of ARVC. Desmoglein 2 (Dsg2)mut/mut mice were used as the ARVC model to verify the therapeutic effects of pharmacological intervention on identified cellular cluster. RESULTS: Right ventricle of ARVC was enriched of CCL3+ proinflammatory macrophages and TNMD+ fibroblasts. Fibroblasts were preferentially affected in ARVC and perturbations associated with ARVC overlap with those reside in genetic variants associated with cardiac arrhythmia. Proinflammatory macrophages strongly interact with fibroblast. Pharmacological inhibition of Nod-like receptor protein 3 (NLRP3), a transcriptional factor predominantly expressed by the CCL3+ proinflammatory macrophages and several other myeloid subclusters, could significantly alleviate right ventricular dilation and dysfunction in Dsg2mut/mut mice (an ARVC mouse model). CONCLUSIONS: This study provided a comprehensive analysis of the lineage-specific changes in the blood and myocardium from ARVC patients at a single-cell resolution. Pharmacological inhibition of NLRP3 could prevent right ventricular dilation and dysfunction of mice with ARVC.

The significance of dynamic monitoring plasma TMAO level in pulmonary arterial hypertension - a cohort study.

BACKGROUND: Gut microbiota assumes an essential role in the development and progression of pulmonary arterial hypertension (PAH). Trimethylamine N-oxide (TMAO), a gut microbiota-dependent metabolite, is correlated with the prognosis of patients with PAH. However, the correlation between changes in TMAO (ΔTMAO) and the prognosis of PAH remains elusive. OBJECTIVES: To investigate the association between ΔTMAO and prognosis of PAH, and explore whether dynamic assessment of TMAO level was superior to measurement at a single time point in predicting prognosis. DESIGN: Single-center cohort study. METHODS: Consecutive patients diagnosed with PAH and had at least two TMAO measurements taken from May 2019 to June 2020 were eligible. The outcome events of this study were defined as adverse clinical events. RESULTS: A total of 117 patients with PAH who had two TMAO measurements and follow-up were included in this study. Patients with ΔTMAO ⩾1.082 µmol/L had over four times increased risk of adverse clinical events than their counterparts after adjusting for confounders [hazard ratio (HR) 4.050, 95% confidence interval (CI): 1.468-11.174; p = 0.007]. Patients with constant high TMAO levels at both time points had the highest risk of adverse clinical events compared with patients with constant low TMAO levels (HR 3.717, 95% CI: 1.627-8.492; p = 0.002). ΔTMAO was also associated with changes in parameters reflecting PAH severity (p < 0.05). CONCLUSION: Changes in TMAO were independently correlated with prognosis in patients with PAH, irrespective of baseline level of TMAO. ΔTMAO also correlated with alteration in disease severity. Repeated assessment of TMAO level contributes to better identification of patients with increased risk of adverse clinical events.

Single-Cell RNA Sequencing of Coronary Perivascular Adipose Tissue From End-Stage Heart Failure Patients Identifies SPP1+ Macrophage Subpopulation as a Target for Alleviating Fibrosis.

BACKGROUND: Perivascular adipose tissue (PVAT) is vital for vascular homeostasis, and PVAT dysfunction is associated with increased atherosclerotic plaque burden. But the mechanisms underlining coronary PVAT dysfunction in coronary atherosclerosis remain elusive. METHODS: We performed single-cell RNA sequencing of the stromal vascular fraction of coronary PVAT from 3 groups of heart transplant recipients with end-stage heart failure, including 3 patients with nonobstructive coronary atherosclerosis, 3 patients with obstructive coronary artery atherosclerosis, and 4 nonatherosclerosis control subjects. Bioinformatics was used to annotate the cellular populations, depict the cellular developmental trajectories and interactions, and explore the differences among 3 groups of coronary PVAT at the cellular and molecular levels. Pathological staining, quantitative real-time polymerase chain reaction, and in vitro studies were performed to validate the key findings. RESULTS: Ten cell types were identified among 67 936 cells from human coronary PVAT. Several cellular subpopulations, including SPP1+ (secreted phosphoprotein 1) macrophages and profibrotic fibroadipogenic progenitor cells, were accumulated in PVAT surrounding atherosclerotic coronary arteries compared with nonatherosclerosis coronary arteries. The fibrosis percentage was increased in PVAT surrounding atherosclerotic coronary arteries, and it was positively associated with the grade of coronary artery stenosis. Cellular interaction analysis suggested OPN (osteopontin) secreted by SPP1+ macrophages interacted with CD44 (cluster of differentiation 44)/integrin on fibroadipogenic progenitor cells. Strikingly, correlation analyses uncovered that higher level of SPP1 in PVAT correlates with a more severe fibrosis degree and a higher coronary stenosis grade. In vitro studies showed that conditioned medium from atherosclerotic coronary PVAT promoted the migration and proliferation of fibroadipogenic progenitor cells, while such effect was prevented by blocking CD44 or integrin. CONCLUSIONS: SPP1+ macrophages accumulated in the PVAT surrounding atherosclerotic coronary arteries, and they promoted the migration and proliferation of fibroadipogenic progenitor cells via OPN-CD44/integrin interaction and thus aggravated the fibrosis of coronary PVAT, which was positively correlated to the coronary stenosis burden. Therefore, SPP1+ macrophages in coronary PVAT may participate in the progression of coronary atherosclerosis.

Alerting trends in epidemiology for calcific aortic valve disease, 1990-2019: An age-period-cohort analysis for the Global Burden of Disease Study 2019.

AIMS: To assess the trends in calcific aortic valve disease (CAVD) epidemiology, with an emphasis on CAVD mortality, leading risk factors, and their associations with age, period, and birth cohort. METHODS AND RESULTS: Prevalence, disability-adjusted life years, and mortality were derived from the Global Burden of Disease Study 2019. The age-period-cohort model was employed to study the detailed trends of CAVD mortality and its leading risk factors. Globally, CAVD showed unsatisfactory results from 1990 to 2019, with the CAVD deaths of 127 000 in 2019. CAVD mortality was substantially reduced in high socio-demographic index (SDI) countries [-1.45%, 95% confidence interval (CI) (-1.61 to -1.30)], mildly increased in high-middle SDI countries [0.22%, 95% CI (0.06-0.37)], and unchanged in other SDI quintiles. There was a noticeable transition in CAVD deaths from younger to older populations globally. The CAVD mortality increased exponentially with age, and the male had higher mortality than the female before 80 years old. Favourable period [0.69, 95% CI (0.66-0.72)] and birth effects [0.30, 95% CI (0.22-0.43)] were mainly observed in high SDI countries, while unfavourable effects were mostly noticed in high-middle SDI countries. High systolic blood pressure was the leading risk factor of CAVD deaths globally, and it showed favourable trends in high SDI regions. CONCLUSION: Although CAVD mortality reduction was observed globally, unfavourable period, and cohort effects were found in many countries. Increase of mortality rate among the population ≥85 years was the common challenge across all SDI quintiles, stressing the necessity to further improve health care for CAVD patients worldwide.

Cellular Landscapes of Nondiseased Human Cardiac Valves From End-Stage Heart Failure-Explanted Heart.

BACKGROUND: Exploring the mechanisms of valvular heart disease at the cellular level may be useful to identify new therapeutic targets; however, the comprehensive cellular landscape of nondiseased human cardiac valve leaflets remains unclear. METHODS: The cellular landscapes of nondiseased human cardiac valve leaflets (5 aortic valves, 5 pulmonary valves, 5 tricuspid valves, and 3 mitral valves) from end-stage heart failure patients undergoing heart transplantation were explored using single-cell RNA sequencing. Bioinformatics was used to identify the cell types, describe the cell functions, and investigate cellular developmental trajectories and interactions. Differences among the 4 types of cardiac valves at the cellular level were summarized. Pathological staining was performed to validate the key findings of single-cell RNA sequencing. An integrative analysis of our single-cell data and published genome-wide association study-based and bulk RNA sequencing-based data provided insights into the cell-specific contributions to calcific aortic valve diseases. RESULTS: Six cell types were identified among 128 412 cells from nondiseased human cardiac valve leaflets. Valvular interstitial cells were the largest population, followed by myeloid cells, lymphocytes, valvular endothelial cells, mast cells, and myofibroblasts. The 4 types of cardiac valve had distinct cellular compositions. The intercellular communication analysis revealed that valvular interstitial cells were at the center of the communication network. The integrative analysis of our single-cell RNA sequencing data revealed key cellular subpopulations involved in the pathogenesis of calcific aortic valve diseases. CONCLUSIONS: The cellular landscape differed among the 4 types of nondiseased cardiac valve, which might explain their differences in susceptibility to pathological remodeling and valvular heart disease.

Cardiac xenotransplantation: a promising way to treat advanced heart failure.

Cardiac xenotransplantation (CXTx) might be a promising approach to bridge the gap between the supply and demand of a donor heart. The survival of cardiac xenograft has been significantly extended in pig-to-nonhuman primate (NHP) CXTx, with records of 195 days and 945 days for orthotropic and heterotopic CXTx, respectively. To present the history of CXTx, we list the reported clinical CXTx, compare pigs and NHPs as sources of hearts, and compare three different kinds of preclinical CXTx models. The application of genetically modified pigs and novel immunosuppressive drugs accelerates the development of CXTx, and we summarize the reported pig-to-NHP CXTx with detailed information. Besides, we discuss the underlining mechanisms and potential preventive strategies of immunological barriers, including hyperacute rejection, acute humoral xenograft rejection, acute cellular xenograft rejection, chronic rejection, coagulation dysfunction, and systemic inflammation. Though intense cellular infiltration in cardiac xenograft has only been documented in a small number of studies, we especially stress the importance of cellular rejection in CXTx, because we believe it is often masked by the rapid and strong humoral response and it may eventually become a more important and common type of xenograft rejection. In addition, we conclude other obstacles as well as possible solutions in CXTx, such as perioperative cardiac xenograft dysfunction, detrimental xenograft overgrowth, and porcine endogenous retroviruses. Finally, we briefly introduce several other approaches that have been proposed to deal with the organ heart shortage crisis, and we firmly believe that CXTx provides the best near-term solution.

Plasma Metabolites-Based Prediction in Cardiac Surgery-Associated Acute Kidney Injury.

Background Cardiac surgery-associated acute kidney injury (CSA-AKI) is a common postoperative complication following cardiac surgery. Currently, there are no reliable methods for the early prediction of CSA-AKI in hospitalized patients. This study developed and evaluated the diagnostic use of metabolomics-based biomarkers in patients with CSA-AKI. Methods and Results A total of 214 individuals (122 patients with acute kidney injury [AKI], 92 patients without AKI as controls) were enrolled in this study. Plasma samples were analyzed by liquid chromatography tandem mass spectrometry using untargeted and targeted metabolomic approaches. Time-dependent effects of selected metabolites were investigated in an AKI swine model. Multiple machine learning algorithms were used to identify plasma metabolites positively associated with CSA-AKI. Metabolomic analyses from plasma samples taken within 24 hours following cardiac surgery were useful for distinguishing patients with AKI from controls without AKI. Gluconic acid, fumaric acid, and pseudouridine were significantly upregulated in patients with AKI. A random forest model constructed with selected clinical parameters and metabolites exhibited excellent discriminative ability (area under curve, 0.939; 95% CI, 0.879-0.998). In the AKI swine model, plasma levels of the 3 discriminating metabolites increased in a time-dependent manner (R2, 0.480-0.945). Use of this AKI predictive model was then confirmed in the validation cohort (area under curve, 0.972; 95% CI, 0.947-0.996). The predictive model remained robust when tested in a subset of patients with early-stage AKI in the validation cohort (area under curve, 0.943; 95% CI, 0.883-1.000). Conclusions High-resolution metabolomics is sufficiently powerful for developing novel biomarkers. Plasma levels of 3 metabolites were useful for the early identification of CSA-AKI.

Metabolic profile of heart tissue in cyanotic congenital heart disease.

BACKGROUND: Cyanotic congenital heart disease (CCHD) is one of the most common birth anomalies, in which chronic hypoxia is the basic pathophysiological process. METHODS: To investigate the heart's metabolic remodeling to hypoxia, we performed an untargeted metabolomic analysis of cardiac tissue from 20 CCHD patients and 15 patients with acyanotic congenital heart disease (ACHD). RESULTS: A total of 71 (63%) metabolites from 113 detected substances in cardiac tissue differed between the CCHD and ACHD groups. A partial least squares discriminant analysis showed separation between the CCHD and ACHD groups. A pathway enrichment analysis revealed that the most enriched metabolic pathways were amino acid metabolism and energy metabolism. Eleven amino acids were increased in CCHD patients, indicating that protein synthesis was down-regulated. Most of the metabolites in Krebs circle were increased in CCHD patients, suggesting down regulation of aerobic energy metabolism. Hierarchical cluster analysis showed that nicotinamide adenine dinucleotide (NAD) was clustered with Krebs cycle related substrates and its level was significantly higher in CCHD than that in ACHD patients. These analyses suggest that NAD might play an important role in response to hypoxia in CCHD patients. CONCLUSION: Our data showed a significantly different metabolic profile in CCHD patients compared to ACHD patients, including reduced protein synthesis and aerobic energy production, and the increased level of NAD in the myocardium may be a response mechanism to hypoxia.

Clinical Application of Machine Learning-Based Artificial Intelligence in the Diagnosis, Prediction, and Classification of Cardiovascular Diseases.

With the rapid development of artificial intelligence (AI) and machine learning (ML), as well as the arrival of the big data era, technological innovations have occurred in the field of cardiovascular medicine. First, the diagnosis of cardiovascular diseases (CVDs) is highly dependent on assistive examinations, the interpretation of which is time consuming and often limited by the knowledge level and clinical experience of doctors; however, AI could be used to automatically interpret the images obtained in auxiliary examinations. Second, some of the predictions of the incidence and prognosis of CVDs are limited in clinical practice by the use of traditional prediction models, but there may be occasions when AI-based prediction models perform well by using ML algorithms. Third, AI has been used to assist precise classification of CVDs by integrating a variety of medical data from patients, which helps better characterize the subgroups of heterogeneous diseases. To help clinicians better understand the applications of AI in CVDs, this review summarizes studies relating to AI-based diagnosis, prediction, and classification of CVDs. Finally, we discuss the challenges of applying AI to cardiovascular medicine.

miR-1268a regulates ABCC1 expression to mediate temozolomide resistance in glioblastoma.

INTRODUCTION: Temozolomide (TMZ) is the preferred chemotherapeutic drug approved for the Glioblastoma multiforme (GBM) treatment. However, resistance to TMZ is the most intractable challenge for treatment of GBM. Screening of miRNAs is becoming a novel strategy to reveal underlying mechanism of drug-resistance of human tumors. MATERIALS AND METHODS: We conducted RNA sequencing (RNA-seq) for GBM cells treated continuously with TMZ 1 or 2 week or not. Bioinformatic analysis was used to predict targets of these altered miRNAs. Subsequently, we studied the potential role of miR-1268a in TMZ-resistance of GBM cells. RESULTS: Expression levels of 55 miRNAs were identified altering both after 1 and 2 weeks TMZ treatment. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were conducted to illuminate the biological implication and related pathways of predicted target genes. We showed that miR-1268a was downregulated after TMZ treatment and targeted ABCC1/MRP1, a membrane transporter contributing to drug resistance, using dual-luciferase assay. Furthermore, we confirmed overexpression of miR-1268a inhibited protein translation of ABCC1 and restored upregulated expression of ABCC1 due to TMZ. Inversely, knockdown of miR-1268a increased ABCC1 at protein level and enhanced upregulation of ABCC1 with TMZ treatment. In addition, our data indicated that miR-1268a enhanced TMZ sensitivity in GBM cells. CONCLUSION: Through RNA-seq analysis, we discovered miR-1268a and elucidated its role in modulating TMZ-resistance of GBM cells by targeting ABCC1.

[Primary culture of human malignant meningioma cells and its intracranial orthotopic transplantation in nude mice].

OBJECTIVE: To obtain stable primary cultures of human malignant meningioma cells and establish an intracranial in-situ tumor model in nude mice. METHODS: Ten surgical specimens of highly suspected malignant meningioma were obtained with postoperative pathological confirmation. Primary malignant meningioma cells were cultured from the tissues using a modified method and passaged. After identification with cell immunofluorescence, the cultured cells were inoculated into the right parietal lobe of 6 nude mice using stereotaxic apparatus and also transplanted subcutaneously in another 6 nude mice. The nude mice were executed after 6 weeks, and HE staining and immunohistochmistry were used to detect tumor growth and the invasion of the adjacent brain tissues. RESULTS: The primary malignant meningioma cells were cultured successfully, and postoperative pathology reported anaplastic malignant meningioma. Cell immunofluorescence revealed positivity for vimentin and EMA in the cells, which showed a S-shaped growth curve in culture. Flow cytometry revealed a cell percentage in the Q3 area of (95.99∓2.58)%. Six weeks after transplantation, tumor nodules occurred in the subcutaneous tumor group, and the nude mice bearing the in situ tumor showed obvious body weight loss. The xenografts in both groups contained a mean of (36∓5.35)% cells expressing Ki-67, and the intracranial in situ tumor showed obvious invasion of the adjacent peripheral brain tissues. CONCLUSION: We obtained stable primary cultures of malignant meningioma cells and successfully established a nude mouse model bearing in situ human malignant meningioma.