Silencing of METTL3 suppressed ferroptosis of myocardial cells by m6A modification of SLC7A11 in a YTHDF2 manner.

Myocardial infarction (MI) is the main cause of heart failure (HF). N6-methyladenosine (m6A) methylation is associated with the progression of HF. The study aimed to explore whether METTL3 regulates ferroptosis of cardiomyocytes in HF. We evaluated ferroptosis by detecting lactic dehydrogenase (LDH) release, lipid reactive oxygen species (ROS), Fe2+, glutathione (GSH), and malonaldehyde (MDA) levels. M6A methylation was assessed using methylated RNA immunoprecipitation assay. The binding relationship was assessed using RNA immunoprecipitation assays. The mRNA stability was assessed using actinomycin D treatment. The results showed that METTL3 was upregulated in oxygen glucose deprivation/recovery (OGD/R) cells, which knockdown suppressed OGD/R-induced ferroptosis. Moreover, METTL3 could bind to SLC7A11, promoting m6A methylation of SLC7A11. Silencing of SLC7A11 abrogated the suppression of ferroptosis induced by METTL3 knockdown. Additionally, YTHDF2 was the reader that recognized the methylation of SLC7A11, reducing the stability of SLC7A11. The silencing of METTL3 inhibited OGD/R-induced ferroptosis by suppressing the m6A methylation of SLC7A11, which is recognized by YTHDF2. The findings suggested that METTL3-mediated ferroptosis might be a new strategy for MI-induced HF therapy.

Toxicity Mechanisms of Nanoplastics on Crop Growth, Interference of Phyllosphere Microbes, and Evidence for Foliar Penetration and Translocation.

Despite the increasing prevalence of atmospheric nanoplastics (NPs), there remains limited research on their phytotoxicity, foliar absorption, and translocation in plants. In this study, we aimed to fill this knowledge gap by investigating the physiological effects of tomato leaves exposed to differently charged NPs and foliar absorption and translocation of NPs. We found that positively charged NPs caused more pronounced physiological effects, including growth inhibition, increased antioxidant enzyme activity, and altered gene expression and metabolite composition and even significantly changed the structure and composition of the phyllosphere microbial community. Also, differently charged NPs exhibited differential foliar absorption and translocation, with the positively charged NPs penetrating more into the leaves and dispersing uniformly within the mesophyll cells. Additionally, NPs absorbed by the leaves were able to translocate to the roots. These findings provide important insights into the interactions between atmospheric NPs and crop plants and demonstrate that NPs' accumulation in crops could negatively impact agricultural production and food safety.

Inhibition of YTHDF1 prevents hypoxia-induced pulmonary artery smooth muscle cell proliferation by regulating Foxm1 translation in an m6A-dependent manner.

Pulmonary arterial hypertension (PAH) is a chronic disease characterized by pulmonary vascular remodeling. It refers to the abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs), and hypoxia is an important risk factor for this progression. The present study aims to investigate the role of YTHDF1 in the regulation of hypoxic PASMC proliferation and the underlying mechanism. Human PASMCs were transfected with si-YTHDF1/2/3 followed by treatment of hypoxia, and the PASMC proliferation and Foxm1 expression were detected. Through RNA pull-down, RNA immunoprecipitation, and protein synthesis assay, the mechanism of YTHDF1 regulating Foxm1 was explored. Next, Foxm1 was inhibited by thiostrepton, and cell proliferation was detected. In vivo, mice received a tail vein injection of adenovirus containing si-YTHDF1 and were exposed to hypoxia treatment. Pulmonary vascular changes, right ventricular systolic pressure (RVSP), and genes involving proliferation were analyzed. YTHDF1 silencing reduced more hypoxic PASMC proliferation and Foxm1 protein level than YTHDF2/3 silencing. Mechanical results showed that YTHDF1 interacted with Foxm1 mRNA and up-regulated Foxm1 protein level by enhancing the translation efficiency in an m6A-dependent manner. Furthermore, YTHDF1 facilitated hypoxic PASMC proliferation and proliferation marker expressions through up-regulation of Foxm1 in an m6A-dependent manner. In vivo, the YTHDF1 silencing alleviated pulmonary vascular changes and fibrosis, reduced RVSP, inhibited the interaction of YTHDF1 and Foxm1, and reduced proliferation marker levels, as compared to the PAH group. In conclusion, YTHDF1 silencing inhibits hypoxic PASMC proliferation by regulating Foxm1 translation in an m6A-dependent manner.

Interfering with Dusp2 alleviates high glucose-induced vascular endothelial cell dysfunction by promoting p38 MAPK pathway activation.

BACKGROUND: Hyperglycemia-induced vascular endothelial cell dysfunction is a major factor contributing to diabetic lower extremity ischemia. We intend to investigate the role of Dusp2 in hyperglycemia-induced vascular endothelial cell dysfunction and related mechanisms. METHODS: The human umbilical vein endothelial cells (HUVECs) were treated with high glucose (HG) as the cell model. Streptozotocin injection was performed to induce diabetes and femoral artery ligation was to induce hind limb ischemia in mice. The levels of Dusp2, p-p38 MAPK, E2F4, and p38 MAPK were evaluated by Western blot or quantitative real-time PCR. The laser Doppler perfusion imaging was conducted to measure blood flow recovery. The cell counting kit-8, transwell, and tube formation assay were performed to evaluate cell proliferation, migration, and angiogenesis, respectively. CD31 immunohistochemical staining was carried out to detect the capillary density of gastrocnemius. The dual-luciferase reporter gene assay and Chromatin immunoprecipitation assay were executed to explore the interaction between E2F4 and Dusp2. RESULTS: Dusp2 was highly expressed in HG-induced HUVECs and diabetic lower extremity ischemia model mice. Interference with Dusp2 promoted cell proliferation, migration, and angiogenesis, as well as alleviated mouse diabetic hindlimb ischemia. Dusp2 knockdown up-regulated p-p38 MAPK levels. We verified the binding between E2F4 and Dusp2. Overexpressing E2F4 suppressed Dusp2 levels and promoted cell proliferation, migration, and angiogenesis, co-overexpression of Dusp2 reversed the results. CONCLUSIONS: Overexpressing E2F4 promotes endothelial cell proliferation, migration, and angiogenesis by inhibiting Dusp2 expression and activating p38 MAPK to alleviate vascular endothelial cell dysfunction under HG stimulation.

The intra-S phase checkpoint directly regulates replication elongation to preserve the integrity of stalled replisomes.

DNA replication is dramatically slowed down under replication stress. The regulation of replication speed is a conserved response in eukaryotes and, in fission yeast, requires the checkpoint kinases Rad3ATR and Cds1Chk2 However, the underlying mechanism of this checkpoint regulation remains unresolved. Here, we report that the Rad3ATR-Cds1Chk2 checkpoint directly targets the Cdc45-MCM-GINS (CMG) replicative helicase under replication stress. When replication forks stall, the Cds1Chk2 kinase directly phosphorylates Cdc45 on the S275, S322, and S397 residues, which significantly reduces CMG helicase activity. Furthermore, in cds1Chk2 -mutated cells, the CMG helicase and DNA polymerases are physically separated, potentially disrupting replisomes and collapsing replication forks. This study demonstrates that the intra-S phase checkpoint directly regulates replication elongation, reduces CMG helicase processivity, prevents CMG helicase delinking from DNA polymerases, and therefore helps preserve the integrity of stalled replisomes and replication forks.

Deficiency of the novel high mobility group protein HMGXB4 protects against systemic inflammation-induced endotoxemia in mice.

Sepsis is a major cause of mortality in intensive care units, which results from a severely dysregulated inflammatory response that ultimately leads to organ failure. While antibiotics can help in the early stages, effective strategies to curtail inflammation remain limited. The high mobility group (HMG) proteins are chromosomal proteins with important roles in regulating gene transcription. While HMGB1 has been shown to play a role in sepsis, the role of other family members including HMGXB4 remains unknown. We found that expression of HMGXB4 is strongly induced in response to lipopolysaccharide (LPS)-elicited inflammation in murine peritoneal macrophages. Genetic deletion of Hmgxb4 protected against LPS-induced lung injury and lethality and cecal ligation and puncture (CLP)-induced lethality in mice, and attenuated LPS-induced proinflammatory gene expression in cultured macrophages. By integrating genome-wide transcriptome profiling and a publicly available ChIP-seq dataset, we identified HMGXB4 as a transcriptional activator that regulates the expression of the proinflammatory gene, Nos2 (inducible nitric oxide synthase 2) by binding to its promoter region, leading to NOS2 induction and excessive NO production and tissue damage. Similar to Hmgxb4 ablation in mice, administration of a pharmacological inhibitor of NOS2 robustly decreased LPS-induced pulmonary vascular permeability and lethality in mice. Additionally, we identified the cell adhesion molecule, ICAM1, as a target of HMGXB4 in endothelial cells that facilitates inflammation by promoting monocyte attachment. In summary, our study reveals a critical role of HMGXB4 in exacerbating endotoxemia via transcriptional induction of Nos2 and Icam1 gene expression and thus targeting HMGXB4 may be an effective therapeutic strategy for the treatment of sepsis.

Atorvastatin-induced tolerogenic dendritic cells improve cardiac remodeling by suppressing TLR-4/NF-κB activation after myocardial infarction.

OBJECTIVE: Myocardial infarction (MI) caused by ischemic cardiomyocyte necrosis induces inflammatory responses that strongly affect ventricular remodeling. Tolerogenic dendritic cells (tDCs) can suppress this effect on inflammatory responses. However, the precise role of atorvastatin-induced tDCs in ventricular remodeling after MI remains unclear. METHODS: To explore the effect of necrotic cardiomyocytes (SNC) and/or atorvastatin on DC function, the expression of CD40, CD80, CD86, and MHC-II was determined using flow cytometry. The protein levels of TLR-4/NF-κB-related molecules were evaluated using western blotting. The infarct area after MI was determined via 2,3,5-triphenyltetrazolium chloride staining. The TUNEL assay was employed to evaluate the apoptosis of cardiomyocytes in heart sections. Masson's trichrome method was used to determine the extent of fibrosis. RESULTS: Compared to the DCs co-cultured with PBS (control), cells co-cultured with Supernatant-IM or Supernatant-NH produced higher levels of inflammatory cytokines, including TNF-α, IL-1, IL-6, IL-12P40, and IL-8. This cytokine production was impaired by atorvastatin treatment. SNC treatment induced DC maturation and enhanced inflammatory cytokine secretion and oxidative stress through TLR-4/NF-κB pathway activation. Compared to that in the PBS-treated group, the left ventricular ejection fraction was significantly improved after tDC treatment. Additionally, compared to that in the PBS-treated group, tDC treatment reduced the left ventricular end-diastolic and end-systolic diameters in mice. Furthermore, treatment with tDCs improved the left ventricular systolic function, attenuated inflammatory cell infiltration, and reduced cardiomyocyte apoptosis, myocardial fibrosis, and infarct size compared to those in the control group. CONCLUSIONS: Adoptive transfer of atorvastatin-induced tDCs alleviated post-infarction cardiomyocyte apoptosis and myocardial fibrosis in association with decreased inflammatory cell infiltration and inhibited oxidative stress, likely by suppressing TLR-4/NF-κB activation after myocardial infarction.

CARMN Is an Evolutionarily Conserved Smooth Muscle Cell-Specific LncRNA That Maintains Contractile Phenotype by Binding Myocardin.

BACKGROUND: Vascular homeostasis is maintained by the differentiated phenotype of vascular smooth muscle cells (VSMCs). The landscape of protein coding genes comprising the transcriptome of differentiated VSMCs has been intensively investigated but many gaps remain including the emerging roles of noncoding genes. METHODS: We reanalyzed large-scale, publicly available bulk and single-cell RNA sequencing datasets from multiple tissues and cell types to identify VSMC-enriched long noncoding RNAs. The in vivo expression pattern of a novel smooth muscle cell (SMC)-expressed long noncoding RNA, Carmn (cardiac mesoderm enhancer-associated noncoding RNA), was investigated using a novel Carmn green fluorescent protein knock-in reporter mouse model. Bioinformatics and quantitative real-time polymerase chain reaction analysis were used to assess CARMN expression changes during VSMC phenotypic modulation in human and murine vascular disease models. In vitro, functional assays were performed by knocking down CARMN with antisense oligonucleotides and overexpressing Carmn by adenovirus in human coronary artery SMCs. Carotid artery injury was performed in SMC-specific Carmn knockout mice to assess neointima formation and the therapeutic potential of reversing CARMN loss was tested in a rat carotid artery balloon injury model. The molecular mechanisms underlying CARMN function were investigated using RNA pull-down, RNA immunoprecipitation, and luciferase reporter assays. RESULTS: We identified CARMN, which was initially annotated as the host gene of the MIR143/145 cluster and recently reported to play a role in cardiac differentiation, as a highly abundant and conserved, SMC-specific long noncoding RNA. Analysis of the Carmn GFP knock-in mouse model confirmed that Carmn is transiently expressed in embryonic cardiomyocytes and thereafter becomes restricted to SMCs. We also found that Carmn is transcribed independently of Mir143/145. CARMN expression is dramatically decreased by vascular disease in humans and murine models and regulates the contractile phenotype of VSMCs in vitro. In vivo, SMC-specific deletion of Carmn significantly exacerbated, whereas overexpression of Carmn markedly attenuated, injury-induced neointima formation in mouse and rat, respectively. Mechanistically, we found that Carmn physically binds to the key transcriptional cofactor myocardin, facilitating its activity and thereby maintaining the contractile phenotype of VSMCs. CONCLUSIONS: CARMN is an evolutionarily conserved SMC-specific long noncoding RNA with a previously unappreciated role in maintaining the contractile phenotype of VSMCs and is the first noncoding RNA discovered to interact with myocardin.

PCSK9 inhibitor recaticimab for hypercholesterolemia on stable statin dose: a randomized, double-blind, placebo-controlled phase 1b/2 study.

BACKGROUND: Recaticimab (SHR-1209, a humanized monoclonal antibody against PCSK9) showed robust LDL-C reduction in healthy volunteers. This study aimed to further assess the efficacy and safety of recaticimab in patients with hypercholesterolemia. METHODS: In this randomized, double-blind, placebo-controlled phase 1b/2 trial, patients receiving stable dose of atorvastatin with an LDL-C level of 2.6 mmol/L or higher were randomized in a ratio of 5:1 to subcutaneous injections of recaticimab or placebo at different doses and schedules. Patients were recruited in the order of 75 mg every 4 weeks (75Q4W), 150Q8W, 300Q12W, 150Q4W, 300Q8W, and 450Q12W. The primary endpoint was percentage change in LDL-C from the baseline to end of treatment (i.e., at week 16 for Q4W and Q8W schedule and at week 24 for Q12W schedule). RESULTS: A total of 91 patients were enrolled and received recaticimab and 19 received placebo. The dose of background atorvastatin in all 110 patients was 10 or 20 mg/day. The main baseline LDL-C ranged from 3.360 to 3.759 mmol/L. The least-squares mean percentage reductions in LDL-C from baseline to end of treatment relative to placebo for recaticimab groups at different doses and schedules ranged from -48.37 to -59.51%. No serious treatment-emergent adverse events (TEAEs) occurred. The most common TEAEs included upper respiratory tract infection, increased alanine aminotransferase, increased blood glucose, and increased gamma-glutamyltransferase. CONCLUSION: Recaticimab as add-on to moderate-intensity statin therapy significantly and substantially reduced the LDL-C level with an infrequent administration schedule (even given once every 12 weeks), compared with placebo. TRIAL REGISTRATION: ClinicalTrials.gov , number NCT03944109.

Long noncoding RNA CASC2 protect ROS-induced oxidative stress in myocardial infarction by miR-18a/SIRT2.

We aimed to investigate the function and its possible mechanisms of long noncoding RNA (lncRNA) in acute myocardial infarction (AMI) model. Patients with AMI and normal volunteers were selected from our hospital. Sprague-Dawley rats were induced into in vivo model of AMI. H9c2 cells were treated with H2 O2 to generate injury model. A significantly lower serum gene expression of lncRNA CASC2 was detected. In rat models of AMI, lncRNA CASC2 gene expressions in heart tissue of mice with AMI were decreased. In in vitro model, downregulation of lncRNA CASC2 increased reactive oxygen species (ROS)-induced oxidative stress; lncRNA CASC2 induced NADPH oxidase (NOX-2) expression and suppressed miR-18a expression; MiR-18a promoted ROS-induced oxidative stress; downregulation of miR-18a decreased ROS-induced oxidative stress. The inhibition of miR-18a reversed the effects of CASC2 downregulation on ROS-induced oxidative stress in in vitro model of AMI. The activation of miR-18a reversed the effects of CASC2 on ROS-induced oxidative stress in in vitro model of AMI. These data for the first time suggest that lncRNA CASC2 have better protective effects on AMI, which could reduce oxidative stress through their carried miR-18a and subsequently downregulating the SIRT2/ROS pathway.

Characteristics, goal-oriented treatments and survival of pulmonary arterial hypertension in China: Insights from a national multicentre prospective registry.

BACKGROUND AND OBJECTIVE: Nationally representative reports on the characteristics and long-term survival of pulmonary arterial hypertension (PAH) from developing countries are scarce. The applicability of the current main risk stratifications and the longitudinal changes in goal-oriented treatments have yet to be elucidated in real-world settings. Therefore, we aimed to provide insights into the characteristics, goal-oriented treatments and survival of PAH in China and to explore the applicability of the main risk stratifications in our independent cohort. METHODS: PAH patients were consecutively enrolled from a national prospective multicentre registry. Data on baseline, follow-up re-evaluation and therapeutic changes were collected. RESULTS: A total of 2031 patients were enrolled, with congenital heart disease (CHD)-PAH (45.2%) being the most common aetiology. The mean age was 35 ± 12 years, and 76.2% were females. At baseline, approximately 20% of the patients with intermediate or high risk received combination treatment. At follow-up, approximately half of the re-evaluated patients did not achieve low-risk profiles, and even among patients who received combination therapy at baseline, 4% of them still worsened. The rate of combination therapy increased significantly from 6.7% before 2015 to 35.5% thereafter. The main risk assessment tools demonstrated good performance for predicting survival both at baseline and at follow-up. CONCLUSION: Chinese PAH patients show both similar and distinct features compared to other countries. Current main risk stratifications can significantly discriminate patients at different risk levels. There were still many patients not achieving low-risk profiles at follow-up, indicating more aggressive treatment should be implemented to optimize the goal-oriented treatment strategy.

GFAP (Glial Fibrillary Acidic Protein)-Positive Progenitor Cells Contribute to the Development of Vascular Smooth Muscle Cells and Endothelial Cells-Brief Report.

OBJECTIVE: While GFAP (glial fibrillary acidic protein) is commonly used as a classical marker for astrocytes in the central nervous system, GFAP-expressing progenitor cells give rise to other cell types during development. The goal of this study was to investigate whether GFAP-expressing progenitor cells contribute to the development of vascular cells in major arteries. Approach and Results: To label GFAP-expressing progenitor cells and their progeny, we crossed GFAP promoter-driven Cre recombinase mice (GFAP-Cre) with transgenic mice expressing the Cre-dependent mTmG dual fluorescent reporter gene. Using this genetic fate-mapping approach, here we demonstrate that GFAP-positive progenitor cells contribute to the development of vascular smooth muscle cells in both neural crest- and non-neural crest-derived vascular beds. In addition, GFAP-positive progenitor cells contribute to a subset of endothelial cells in some vasculature. Furthermore, fate-mapping analyses at multiple time points of mouse development demonstrate a time-dependent increase in the contribution of GFAP-positive progenitors to vascular smooth muscle cells, which mostly occurs in the postnatal period. CONCLUSIONS: Our study demonstrates that vascular smooth muscle cells and endothelial cells within the same vascular segment are developmentally heterogeneous, where varying proportions of vascular smooth muscle cells and endothelial cells are contributed by GFAP-positive progenitor cells.

Metformin Attenuates Hypoxia-induced Endothelial Cell Injury by Activating the AMP-Activated Protein Kinase Pathway.

ABSTRACT: Metformin reduces the incidence of cardiovascular diseases, and potential underlying mechanisms of action have been suggested. Here, we investigated the role of metformin in endothelial cell injury and endothelial-mesenchymal transition (EndMT) induced by hypoxia. All experiments were performed in human cardiac microvascular endothelial cells (HCMECs). HCMECs were exposed to hypoxic conditions for 24, 48, 72, and 96 hours, and we assessed the cell viability by cell counting kit 8; metformin (2, 5, 10, and 20 mmol/L) was added to the cells after exposure to the hypoxic conditions for 48 hours. The cells were randomly divided into the control group, hypoxia group, hypoxia + metformin group, hypoxia + control small interfering RNA group, hypoxia + small interfering Prkaa1 (siPrkaa1) group, and hypoxia + siPrkaa1 + metformin group. Flow cytometry and cell counting kit 8 were used to monitor apoptosis and assess cell viability. Immunofluorescence staining was used to identify the CD31+/alpha smooth muscle actin+ double-positive cells. Quantitative real-time-PCR and Western blot were used for mRNA and protein expression analyses, respectively. Hypoxia contributed to endothelial injuries and EndMT of HCMECs in a time-dependent manner, which was mainly manifested as decreases in cell viability, increases in apoptotic rate, and changes in expression of apoptosis-related and EndMT-related mRNAs and proteins. Furthermore, metformin could attenuate the injuries and EndMT caused by hypoxia. After metformin treatment, phosphorylated-AMPK (pAMPK) and p-endothelial nitric oxide synthase expression increased, whereas p-mammalian target of rapamycin expression decreased. However, results obtained after transfection with siPrkaa1 were in contrast to the results of metformin treatment. In conclusion, metformin can attenuate endothelial injuries and suppress EndMT of HCMECs under hypoxic conditions because of its ability to activate the AMPK pathway, increase p-AMPK/AMP-activated protein kinase, and inhibit mammalian target of rapamycin.

Characteristics and long-term survival of patients with chronic thromboembolic pulmonary hypertension in China.

BACKGROUND AND OBJECTIVE: The purpose of this study was to report the characteristics and long-term survival of patients with CTEPH treated in three distinct ways: PEA, BPA and medical therapy. METHODS: Patients diagnosed with CTEPH were included in the registry that was set up in 18 centres from August 2009 to July 2018. The characteristics and survival of patients with CTEPH receiving the different treatments were reported. Prognostic factors were evaluated by Cox regression model. RESULTS: A total of 593 patients with CTEPH were included. Eighty-one patients were treated with PEA, 61 with BPA and 451 with drugs. The estimated survival rates at 1, 3, 5 and 8 years were, respectively, 95.2%, 84.6%, 73.4% and 66.6% in all patients; 92.6%, 89.6%, 87.5% and 80.2% in surgical patients; and 95.4%, 88.3%, 71.0% and 64.1% in medically treated patients. The estimated survival rates at 1, 3, 5 and 7 years in patients treated with BPA were 96.7%, 88.1%, 70.0% and 70.0%, respectively. For all patients, PEA was an independent predictor of survival. Other independent risk factors were CHD, cardiac index, PVR, big endothelin-1, APE and 6MWD. CONCLUSION: This is the first multicentre prospective registry reporting baseline characteristics and estimated survival of patients with CTEPH in China. The long-term survival rates are similar to those of patients in the international and Spanish registries. PEA is an independent predictor of survival.

Risk prediction in medically treated chronic thromboembolic pulmonary hypertension.

BACKGROUND: At present, there is no generally accepted comprehensive prognostic risk prediction model for medically treated chronic thromboembolic pulmonary hypertension (CTEPH) patients. METHODS: Consecutive medically treated CTEPH patients were enrolled in a national multicenter prospective registry study from August 2009 to July 2018. A multivariable Cox proportional hazards model was utilized to derive the prognostic model, and a simplified risk score was created thereafter. Model performance was evaluated in terms of discrimination and calibration, and compared to the Swedish/COMPERA risk stratification method. Internal and external validation were conducted to validate the model performance. RESULTS: A total of 432 patients were enrolled. During a median follow-up time of 38.73 months (IQR: 20.79, 66.10), 94 patients (21.8%) died. The 1-, 3-, and 5-year survival estimates were 95.5%, 83.7%, and 70.9%, respectively. The final model included the following variables: the Swedish/COMPERA risk stratum (low-, intermediate- or high-risk stratum), pulmonary vascular resistance (PVR, ≤ or > 1600 dyn·s/cm5), total bilirubin (TBIL, ≤ or > 38 µmol/L) and chronic kidney disease (CKD, no or yes). Compared with the Swedish/COMPERA risk stratification method alone, both the derived model [C-index: 0.715; net reclassification improvement (NRI): 0.300; integrated discriminatory index (IDI): 0.095] and the risk score (C-index: 0.713; NRI: 0.300; IDI: 0.093) showed improved discriminatory power. The performance was validated in a validation cohort of 84 patients (C-index = 0.707 for the model and 0.721 for the risk score). CONCLUSIONS: A novel risk stratification strategy can serve as a useful tool for determining prognosis and guide management for medically treated CTEPH patients. TRIAL REGISTRATION: ClinicalTrials.gov (Identifier: NCT01417338).

Paroxysmal short runs of irregular narrow QRS tachycardia: What is the mechanism?

The present case that showed the frequent transition between left atrial tachycardia and AV nodal re-entrant tachycardia has never been reported. This instructive case highlights the need for careful interpretation of intracardiac electrocardiogram and suggests that differential pacing maneuvers are not feasible in transitional tachycardia with atrial fusion.

Activation of CaMKII via ER-stress mediates coxsackievirus B3-induced cardiomyocyte apoptosis.

Cardiomyocyte apoptosis contributes to the development of coxsackievirus B3 (CVB3)-induced myocarditis, but the mechanism for the apoptosis by CVB3 infection remains unclear. Here, we showed that CVB3-induced endoplasmic reticulum (ER) stress response and apoptosis in cultured H9c2 cardiomyocytes. We found that Ca2+ -calmodulin-dependent kinase II (CaMKII) was activated by ER stress-dependent intracellular Ca2+ overload in the CVB3-infected H9c2 cardiomyocytes. Treatment with an inhibitor of ER stress, 4-phenylbutyric acid (4-PBA), attenuated intracellular Ca2+ accumulation indirectly and reduced CaMKII activity. Inhibition of CaMKII with pharmacological inhibitor (KN-93) or short hairpin RNA reduced CVB3-induced H9c2 apoptosis and repressed cytochrome c release from mitochondria to cytoplasm; whereas overexpression of the activated mutant of CaMKII (CaMKII-T287D) enhanced CVB3-induced H9c2 apoptosis and mitochondrial cytochrome c release, which could be alleviated by blocking of mitochondrial Ca2+ uniporter or mitochondrial permeability transition pore. Further in vivo investigation revealed that blocking of CaMKII with KN-93 prevented cardiomyocytes apoptosis and improved cardiac contractile function in CVB3-infected mouse heart. Collectively, these findings provide a novel evidence that CaMKII plays a vital role in the promotion of CVB3-induced cardiomyocyte apoptosis, which links ER stress and mitochondrial Ca2+ uptake.

Effects of PGC1α on myocardial ischemia reperfusion injury and the underlying mechanisms. / PGC1αå¯¹å¿ƒè‚Œç¼ºè¡€å†çŒæ³¨æŸä¼¤çš„ä½œç”¨åŠå ¶æœºåˆ¶.

OBJECTIVES: Peroxisome proliferator-activated receptor gamma coactivator 1α (PGC1α) controls mitochondrial biogenesis, but its role in cardiovascular diseases is unclear. The purpose of this study is to explore the effect of PGC1α on myocardial ischemia-reperfusion injury and the underlying mechanisms. METHODS: The transverse coronary artery of SD rat was ligated for 30 minutes followed by 2 hours of reperfusion. Triphenyltetrazolium chloride (TTC) staining was performed to measure the area of myocardial infarction. Immunohistochemistry and Western blotting were used to detect the PGC1α expression in myocardium. The rat cardiomyocyte H9C2 was subjected to hypoxia/reoxygenation (H/R) with the knockdown of PGC1α or hypoxia- inducible factor 1α (HIF-1α), or with treatment of metformin. Western blotting was used to detect the expression of PGC1α, HIF-1α, p21, BAX, and caspase-3. CCK-8 was performed to detect cell viability, and flow cytometry was used to detect apoptosis and mitochondrial superoxide (mitoSOX) release. RT-qPCR was used to detect the mRNA expression of PGC1α and HIF-1α. Besides, chromatin immunoprecipitation (ChIP)-qPCR and luciferase reporter gene assay were applied to detect the transcriptional regulation effect of HIF-1α on PGC1α. RESULTS: After I/R, the PGC1α expression was increased in infarcted myocardium. H/R induced H9C2 cell apoptosis (P<0.001). The release of mitoSOX (P<0.001) and protein expression of PGC1α, and apoptosis-related p21, BAX, and caspase-3 were increased. However, knockdown of PGC1α reduced apoptosis (P<0.01) and mitoSOX release (P<0.001), and decreased protein expression of apoptosis-related gene. HIF-1α is bound to the promoter region of PGC1α. Knockdown of HIF-1α inhibited the transcription and protein expression of PGC1α and further to reduce the apoptosis (all P<0.001) and mitoSOX release (P<0.01). While overexpression of PGC1α reversed the effects caused by HIF-1α knockdown (all P<0.001). Metformin effectively reduced H/R-induced apoptosis (P=0.013), mitoSOX release (P<0.001), HIF-1α, PGC1α and apoptosis-related protein expression, recovered the cell viability (P<0.001), and reduced myocardial infarction (P=0.003). CONCLUSIONS: After I/R, HIF-1α up-regulates the expression of PGC1α, leading to an increase in ROS production and aggravation of injury. Metformin can inhibit the accumulation of HIF-1α during hypoxia and effectively protect myocardium from ischemia/reperfusion injury.

Lovastatin attenuates angiotensin II induced cardiovascular fibrosis through the suppression of YAP/TAZ signaling.

Angiotensin II (ANG II) is associated with fibrosis in both clinical and basic studies. Thus, we aimed to explore a mechanism by which ANG II induces fibrosis. 5 µM of ANG II was used in the in vitro study. The mouse cardiovascular fibrosis model was established by infused with AngII (1000 ng/kg/min) for 7 days and cotreated with lovastatin (10 mg/kg daily) or vehicle control (DMSO in saline). We found that ANG II activated yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ), two transcription factors that were shown to induce fibrosis. Inhibition of ras homolog gene family member A (RhoA) reduced ANG II-induced YAP/TAZ transcriptional activity, which suggests that the upregulation of YAP/TAZ signaling by ANG II is RhoA-dependent. Furthermore, studies have shown that the inhibition of YAP/TAZ by either siRNA or small molecule inhibitor suppressed ANG II-induced expression of fibrogenic genes, indicating that ANG II upregulates YAP/TAZ to initiate fibrosis. The mevalonate pathway, which is targeted by statins, has also been shown to control YAP/TAZ. Here, we found that the suppression of YAP/TAZ signaling by lovastatin attenuates ANG II-induced fibrosis, both in vitro and in vivo. These data reveal a novel mechanism for ANG II in the induction of fibrosis. In addition, our findings provide a reasonable explanation regarding the mechanism by which statins improve fibrosis in patients with cardiovascular and renal diseases.

Fibrillatory wave amplitude on transesophageal ECG as a marker of left atrial low-voltage areas in patients with persistent atrial fibrillation.

BACKGROUND: Low-voltage areas (LVAs) are frequently observed in patients with persistent atrial fibrillation (PeAF) and may represent adverse atrial remodeling. However, noninvasive method of evaluating LAVs is not well established. METHODS: In a cohort of 68 patients with PeAF, endocardial voltage maps of left atrium (LA) were created during sinus rhythm after pulmonary vein isolation (PVI). LVAs were defined as areas with electrogram amplitudes <0.5 mV. LA-LVAs were correlated with clinical, echocardiographic, surface, and transesophageal electrocardiography (TE-ECG) variables. RESULTS: LA voltage mapping revealed any degree of LA-LVAs in 50 (73.5%) patients. Patients with LA-LVAs were older, had a longer history of AF, and lower fibrillatory wave (F wave) amplitude on TE-ECG (0.27 ± 0.06 vs 0.39 ± 0.08 mv, p < .01) as compared to patients without LA-LVAs. The extent of LA-LVAs was weakly correlated with age (R = 0.36, p = .03) and AF duration (R = 0.26, p = .02), but significantly correlated with F-wave amplitude on TE-ECG (R = -0.57, p < .01). Only F-wave amplitude on TE-ECG was found as independent predictor for the presence of LA-LVAs (OR = 1.53, 95% CI = 1.09-2.96, p = .03). A receiver operating characteristic (ROC) curve identified an F-wave amplitude of 0.29 mV (AUC = 0.788; sensitivity = 68.4%; specificity = 73.2%) on TE-ECG as the optimal cutoff value for predicting LA-LVAs. CONCLUSIONS: As a noninvasive investigation, F-wave amplitude on TE-ECG may be used as an indicator for the presence of LA-LVAs.