Mitochondrial GSNOR Alleviates Cardiac Dysfunction via ANT1 Denitrosylation.

BACKGROUND: The cardiac-protective role of GSNOR (S-nitrosoglutathione reductase) in the cytoplasm, as a denitrosylase enzyme of S-nitrosylation, has been reported in cardiac remodeling, but whether GSNOR is localized in other organelles and exerts novel effects remains unknown. We aimed to elucidate the effects of mitochondrial GSNOR, a novel subcellular localization of GSNOR, on cardiac remodeling and heart failure (HF). METHODS: GSNOR subcellular localization was observed by cellular fractionation assay, immunofluorescent staining, and colloidal gold particle staining. Overexpression of GSNOR in mitochondria was achieved by mitochondria-targeting sequence-directed adeno-associated virus 9. Cardiac-specific knockout of GSNOR mice was used to examine the role of GSNOR in HF. S-nitrosylation sites of ANT1 (adenine nucleotide translocase 1) were identified using biotin-switch and liquid chromatography-tandem mass spectrometry. RESULTS: GSNOR expression was suppressed in cardiac tissues of patients with HF. Consistently, cardiac-specific knockout mice showed aggravated pathological remodeling induced by transverse aortic constriction. We found that GSNOR is also localized in mitochondria. In the angiotensin II-induced hypertrophic cardiomyocytes, mitochondrial GSNOR levels significantly decreased along with mitochondrial functional impairment. Restoration of mitochondrial GSNOR levels in cardiac-specific knockout mice significantly improved mitochondrial function and cardiac performance in transverse aortic constriction-induced HF mice. Mechanistically, we identified ANT1 as a direct target of GSNOR. A decrease in mitochondrial GSNOR under HF leads to an elevation of S-nitrosylation ANT1 at cysteine 160 (C160). In accordance with these findings, overexpression of either mitochondrial GSNOR or ANT1 C160A, non-nitrosylated mutant, significantly improved mitochondrial function, maintained the mitochondrial membrane potential, and upregulated mitophagy. CONCLUSIONS: We identified a novel species of GSNOR localized in mitochondria and found mitochondrial GSNOR plays an essential role in maintaining mitochondrial homeostasis through ANT1 denitrosylation, which provides a potential novel therapeutic target for HF.

Ataxia telangiectasia mutated kinase inhibition promotes irradiation-induced PD-L1 expression in tumour-associated macrophages through IFN-I/JAK signalling pathway.

Tumour-associated macrophages (TAMs) are one of the primary sources of PD-L1 expression in the tumour microenvironment (TME). Ionizing radiation (IR) promotes PD-L1 expression in tumour cells. However, the effect of IR on macrophage PD-L1 expression and the underlying mechanisms remain unclear. ATM kinase, as the key kinase for initiating DNA damage repair (DDR) process, is associated with innate immune STING axis activation. Here, we explored the molecular mechanism implicated in macrophage PD-L1 expression regulated by IR as well as the role of ATM kinase in this process. IR-regulated PD-L1 expression in macrophages and associated signalling pathways were explored by in vitro studies using murine and human macrophage cell lines. A colorectal xenograft murine model was employed to demonstrate the impact of targeting ATM and PD-L1 expression in TAMs following IR on growth of tumour in vivo. IR up-regulated PD-L1 expression in macrophages, which was further augmented by ATM kinase inhibition. ATM inhibition increased IR-induced DNA damage, which activated STING/interferon regulatory factor 3 (IRF3) signalling pathway and up-regulated type I interferon (IFN-I) expression in macrophages. IFN-I bound to the IFN α receptor 1 on macrophages, activated the downstream JAK1 and STAT1/3 signalling and eventually led to PD-L1 up-expression. ATM inhibition augmented IR-induced PD-L1 expression in macrophages and CD8+ T cell infiltration, and promoted anti-tumour efficacy in vivo. These results suggested that ATM inhibition promoted IR-induced PD-L1 expression through the activation of innate immunity in TAMs, which provided a novel approach to enhance the anti-tumour efficacy of RT.

HINT1 (Histidine Triad Nucleotide-Binding Protein 1) Attenuates Cardiac Hypertrophy Via Suppressing HOXA5 (Homeobox A5) Expression.

BACKGROUND: Cardiac hypertrophy is an important prepathology of, and will ultimately lead to, heart failure. However, the mechanisms underlying pathological cardiac hypertrophy remain largely unknown. This study aims to elucidate the effects and mechanisms of HINT1 (histidine triad nucleotide-binding protein 1) in cardiac hypertrophy and heart failure. METHODS: HINT1 was downregulated in human hypertrophic heart samples compared with nonhypertrophic samples by mass spectrometry analysis. Hint1 knockout mice were challenged with transverse aortic constriction surgery. Cardiac-specific overexpression of HINT1 mice by intravenous injection of adeno-associated virus 9 (AAV9)-encoding Hint1 under the cTnT (cardiac troponin T) promoter were subjected to transverse aortic construction. Unbiased transcriptional analyses were used to identify the downstream targets of HINT1. AAV9 bearing shRNA against Hoxa5 (homeobox A5) was administrated to investigate whether the effects of HINT1 on cardiac hypertrophy were HOXA5-dependent. RNA sequencing analysis was performed to recapitulate possible changes in transcriptome profile.Coimmunoprecipitation assays and cellular fractionation analyses were conducted to examine the mechanism by which HINT1 regulates the expression of HOXA5. RESULTS: The reduction of HINT1 expression was observed in the hearts of hypertrophic patients and pressure overloaded-induced hypertrophic mice, respectively. In Hint1-deficient mice, cardiac hypertrophy deteriorated after transverse aortic construction. Conversely, cardiac-specific overexpression of HINT1 alleviated cardiac hypertrophy and dysfunction. Unbiased profiler polymerase chain reaction array showed HOXA5 is 1 target for HINT1, and the cardioprotective role of HINT1 was abolished by HOXA5 knockdown in vivo. Hoxa5 was identified to affect hypertrophy through the TGF-ß (transforming growth factor ß) signal pathway. Mechanically, HINT1 inhibited PKCß1 (protein kinase C ß type 1) membrane translocation and phosphorylation via direct interaction, attenuating the MEK/ERK/YY1 (mitogen-activated protein kinase/extracellular signal-regulated kinase kinase/yin yang 1) signal pathway, downregulating HOXA5 expression, and eventually attenuating cardiac hypertrophy. CONCLUSIONS: HINT1 protects against cardiac hypertrophy through suppressing HOXA5 expression. These findings indicate that HINT1 may be a potential target for therapeutic interventions in cardiac hypertrophy and heart failure.

SAMHD1 silencing cooperates with radiotherapy to enhance anti-tumor immunity through IFI16-STING pathway in lung adenocarcinoma.

BACKGROUND: Sterile alpha motif domain and histidine-aspartate domain-containing protein 1 (SAMHD1) is a DNA end resection factor, which is involved in DNA damage repair and innate immunity. However, the role of SAMHD1 in anti-tumor immunity is still unknown. This study investigated the effects of SAMHD1 on stimulator of interferon genes (STING)-type I interferon (IFN) pathway and radiation-induced immune responses. METHODS: The roles of SAMHD1 in the activation of cytosolic DNA sensing STING pathway in lung adenocarcinoma (LUAD) cells were investigated with flow cytometry, immunofluorescence, immunoblotting and qPCR. The combined effects of SAMHD1 silencing and radiation on tumor cell growth and STING pathway activation were also evaluated with colony formation and CCK8 assay. The Lewis lung cancer mouse model was used to evaluate the combined efficiency of SAMHD1 silencing and radiotherapy in vivo. Macrophage M1 polarization and cytotoxic T cell infiltration were evaluated with flow cytometry. RESULTS: The single-stranded DNA (ssDNA) accumulated in the cytosol of SAMHD1-deficient lung adenocarcinoma (LUAD) cells, accompanied by upregulated DNA sensor IFN-γ-inducible protein 16 (IFI16) and activated STING pathway. The translocation of IFI16 from nucleus to cytosol was detected in SAMHD1-deficient cells. IFI16 and STING were acquired in the activation of STING-IFN-I pathway in SAMHD1-deficient cells. SAMHD1 silencing in LUAD cells promoted macrophage M1 polarization in vitro. SAMHD1 silencing synergized with radiation to activate ssDNA-STING-IFN-I pathway, inhibit proliferation, promote apoptosis and regulate cell cycle. SAMHD1 silencing cooperated with radiotherapy to inhibit tumor growth and increase CD86+MHC-IIhigh M1 proportion and CD8+ T cell infiltration in vivo. CONCLUSIONS: SAMHD1 deficiency induced IFN-I production through cytosolic IFI16-STING pathway in LUAD cells. Moreover, SAMHD1 downregulation and radiation cooperated to inhibit tumor growth and enhance anti-tumor immune responses through macrophage M1 polarization and CD8+ T cell infiltration. Combination of SAMHD1 inhibition and radiotherapy may be a potentially therapeutic strategy for LUAD patients.

Prediction of radiosensitivity and radiocurability using a novel supervised artificial neural network.

BACKGROUND: Radiotherapy has been widely used to treat various cancers, but its efficacy depends on the individual involved. Traditional gene-based machine-learning models have been widely used to predict radiosensitivity. However, there is still a lack of emerging powerful models, artificial neural networks (ANN), in the practice of gene-based radiosensitivity prediction. In addition, ANN may overfit and learn biologically irrelevant features. METHODS: We developed a novel ANN with Selective Connection based on Gene Patterns (namely ANN-SCGP) to predict radiosensitivity and radiocurability. We creatively used gene patterns (gene similarity or gene interaction information) to control the "on-off" of the first layer of weights, enabling the low-dimensional features to learn the gene pattern information. ANN-SCGP was trained and tested in 82 cell lines and 1,101 patients from the 11 pan-cancer cohorts. RESULTS: For survival fraction at 2 Gy, the root mean squared errors (RMSE) of prediction in ANN-SCGP was the smallest among all algorithms (mean RMSE: 0.1587-0.1654). For radiocurability, ANN-SCGP achieved the first and second largest C-index in the 12/20 and 4/20 tests, respectively. The low dimensional output of ANN-SCGP reproduced the patterns of gene similarity. Moreover, the pan-cancer analysis indicated that immune signals and DNA damage responses were associated with radiocurability. CONCLUSIONS: As a model including gene pattern information, ANN-SCGP had superior prediction abilities than traditional models. Our work provided novel insights into radiosensitivity and radiocurability.

S-nitrosylation of c-Jun N-terminal kinase mediates pressure overload-induced cardiac dysfunction and fibrosis.

Cardiac fibrosis (CF) is an irreversible pathological process that occurs in almost all kinds of cardiovascular diseases. Phosphorylation-dependent activation of c-Jun N-terminal kinase (JNK) induces cardiac fibrosis. However, whether S-nitrosylation of JNK mediates cardiac fibrosis remains an open question. A biotin-switch assay confirmed that S-nitrosylation of JNK (SNO-JNK) increased significantly in the heart tissues of hypertrophic patients, transverse aortic constriction (TAC) mice, spontaneously hypertensive rats (SHRs), and neonatal rat cardiac fibroblasts (NRCFs) stimulated with angiotensin II (Ang II). Site to site substitution of alanine for cysteine in JNK was applied to determine the S-nitrosylated site. S-Nitrosylation occurred at both Cys116 and Cys163 and substitution of alanine for cysteine 116 and cysteine 163 (C116/163A) inhibited Ang II-induced myofibroblast transformation. We further confirmed that the source of S-nitrosylation was inducible nitric oxide synthase (iNOS). 1400 W, an inhibitor of iNOS, abrogated the profibrotic effects of Ang II in NRCFs. Mechanistically, SNO-JNK facilitated the nuclear translocation of JNK, increased the phosphorylation of c-Jun, and induced the transcriptional activity of AP-1 as determined by chromatin immunoprecipitation and EMSA. Finally, WT and iNOS-/- mice were subjected to TAC and iNOS knockout reduced SNO-JNK and alleviated cardiac fibrosis. Our findings demonstrate an alternative mechanism by which iNOS-induced SNO-JNK increases JNK pathway activity and accelerates cardiac fibrosis. Targeting SNO-JNK might be a novel therapeutic strategy against cardiac fibrosis.

S-nitrosylation of Hsp90 promotes cardiac hypertrophy in mice through GSK3ß signaling.

Cardiac hypertrophy, as one of the major predisposing factors for chronic heart failure, lacks effective interventions. Exploring the pathogenesis of cardiac hypertrophy will reveal potential therapeutic targets. S-nitrosylation is a kind of posttranslational modification that occurs at active cysteines of proteins to mediate various cellular processes. We here identified heat shock protein 90 (Hsp90) as a highly S-nitrosylated target in the hearts of rodents with hypertrophy, and the role of Hsp90 in cardiac hypertrophy remains undefined. The S-nitrosylation of Hsp90 (SNO-Hsp90) levels were elevated in angiotensin II (Ang II)- or phenylephrine (PE)-treated neonatal rat cardiomyocytes (NRCMs) in vitro as well as in cardiomyocytes isolated from mice subjected to transverse aortic constriction (TAC) in vivo. We demonstrated that the elevated SNO-Hsp90 levels were mediated by decreased S-nitrosoglutathione reductase (GSNOR) expression during cardiac hypertrophy, and delivery of GSNOR adeno-associated virus expression vectors (AAV9-GSNOR) decreased the SNO-Hsp90 levels to attenuate cardiac hypertrophy. Mass spectrometry analysis revealed that cysteine 589 (Cys589) might be the S-nitrosylation site of Hsp90. Delivery of the mutated AAV9-Hsp90-C589A inhibited SNO-Hsp90 levels and attenuated cardiac hypertrophy. We further revealed that SNO-Hsp90 led to increased interaction of glycogen synthase kinase 3ß (GSK3ß) and Hsp90, leading to elevated GSK3ß phosphorylation and decreased eIF2Bε phosphorylation, thereby aggravating cardiac hypertrophy. Application of GSK3ß inhibitor TWS119 abolished the protective effect of Hsp90-C589A mutation in Ang II-treated NRCMs. In conclusion, this study demonstrates a critical role of SNO-Hsp90 in cardiac hypertrophy, which may be of a therapeutic target for cardiac hypertrophy treatment.

SASS6 promotes proliferation of esophageal squamous carcinoma cells by inhibiting the p53 signaling pathway.

SASS6 encodes for the Homo sapiens SAS-6 centriolar assembly protein and is important for proper centrosome formation. Although centrosomes are amplified in a wide variety of tumor types, abnormally high SASS6 expression had previously only been identified in colon cancer. Moreover, the role of SASS6 in esophageal squamous cell carcinoma (ESCC) pathogenesis has not yet been elucidated. The aim of this study was to investigate the role and mechanisms of SASS6 in ESCC. In this study, we found that the mRNA and protein levels of SASS6 were increased in human ESCC samples. In addition, SASS6 protein expression was associated with the esophageal cancer stage and negatively affected survival of patients with ESCC. Furthermore, silencing of SASS6 inhibited cell growth and promoted apoptosis of ESCC cells in vitro and inhibited xenograft tumor formation in vivo. A genetic cluster and pathway analysis showed that SASS6 regulated the p53 signaling pathway. Western blot demonstrated that CCND2, GADD45A and EIF4EBP1 protein expression decreased and that TP53 protein expression increased after the knockdown of SASS6 in ESCC cells. Therefore, SASS6 promoted the proliferation of esophageal cancer by inhibiting the p53 signaling pathway. SASS6 has potential as a novel tumor marker and a therapeutic target for ESCC.

An improved procedure for isolating adult mouse cardiomyocytes for epicardial activation mapping.

Cardiovascular disease is a leading cause of death and disability worldwide. Although genetically modified mouse models offer great potential for robust research in vivo, in vitro studies using isolated cardiomyocytes also provide an important approach for investigating the mechanisms underlying cardiovascular disease pathogenesis and drug actions. Currently, isolation of mouse adult cardiomyocytes often relies on aortic retrograde intubation under a stereoscopic microscope, which poses considerable technical barriers and requires extensive training. Although a simplified, Langendorff-free method has been used to isolate viable cardiomyocytes from the adult mouse heart, the system requires enzymatic digestions and continuous manual technical operation. This study established an optimized approach that allows isolation of adult mouse cardiomyocytes and epicardial activation mapping of mouse hearts using a Langendorff device. We used retrograde puncture through the abdominal aorta in vivo and enzymatic digestion on the Langendorff perfusion device to isolate adult mouse cardiomyocytes without using a microscope. The yields of isolated cardiomyocytes were amenable to patch clamp techniques. Furthermore, this approach allowed epicardial activation mapping. We used a novel, simplified method to isolate viable cardiomyocytes from adult mouse hearts and to map epicardial activation. This novel approach could be beneficial in more extensive research in the cardiac field.

SNO-MLP (S-Nitrosylation of Muscle LIM Protein) Facilitates Myocardial Hypertrophy Through TLR3 (Toll-Like Receptor 3)-Mediated RIP3 (Receptor-Interacting Protein Kinase 3) and NLRP3 (NOD-Like Receptor Pyrin Domain Containing 3) Inflammasome Activation.

BACKGROUND: S-nitrosylation (SNO), a prototypic redox-based posttranslational modification, is involved in the pathogenesis of cardiovascular disease. The aim of this study was to determine the role of SNO of MLP (muscle LIM protein) in myocardial hypertrophy, as well as the mechanism by which SNO-MLP modulates hypertrophic growth in response to pressure overload. METHODS: Myocardial samples from patients and animal models exhibiting myocardial hypertrophy were examined for SNO-MLP level using biotin-switch methods. SNO sites were further identified through liquid chromatography-tandem mass spectrometry. Denitrosylation of MLP by the mutation of nitrosylation sites or overexpression of S-nitrosoglutathione reductase was used to analyze the contribution of SNO-MLP in myocardial hypertrophy. Downstream effectors of SNO-MLP were screened through mass spectrometry and confirmed by coimmunoprecipitation. Recruitment of TLR3 (Toll-like receptor 3) by SNO-MLP in myocardial hypertrophy was examined in TLR3 small interfering RNA-transfected neonatal rat cardiomyocytes and in a TLR3 knockout mouse model. RESULTS: SNO-MLP level was significantly higher in hypertrophic myocardium from patients and in spontaneously hypertensive rats and mice subjected to transverse aortic constriction. The level of SNO-MLP also increased in angiotensin II- or phenylephrine-treated neonatal rat cardiomyocytes. S-nitrosylated site of MLP at cysteine 79 was identified by liquid chromatography-tandem mass spectrometry and confirmed in neonatal rat cardiomyocytes. Mutation of cysteine 79 significantly reduced hypertrophic growth in angiotensin II- or phenylephrine-treated neonatal rat cardiomyocytes and transverse aortic constriction mice. Reducing SNO-MLP level by overexpression of S-nitrosoglutathione reductase greatly attenuated myocardial hypertrophy. Mechanistically, SNO-MLP stimulated TLR3 binding to MLP in response to hypertrophic stimuli, and disrupted this interaction by downregulating TLR3-attenuated myocardial hypertrophy. SNO-MLP also increased the complex formation between TLR3 and RIP3 (receptor-interacting protein kinase 3). This interaction in turn induced NLRP3 (nucleotide-binding oligomerization domain-like receptor pyrin domain containing 3) inflammasome activation, thereby promoting the development of myocardial hypertrophy. CONCLUSIONS: Our findings revealed a key role of SNO-MLP in myocardial hypertrophy and demonstrated TLR3-mediated RIP3 and NLRP3 inflammasome activation as the downstream signaling pathway, which may represent a therapeutic target for myocardial hypertrophy and heart failure.

Immune and stromal scoring system associated with tumor microenvironment and prognosis: a gene-based multi-cancer analysis.

BACKGROUND: Tumor microenvironment (TME) is associated with tumor progression and prognosis. Previous studies provided tools to estimate immune and stromal cell infiltration in TME. However, there is still a lack of single index to reflect both immune and stromal status associated with prognosis and immunotherapy responses. METHODS: A novel immune and stromal scoring system named ISTMEscore was developed. A total of 15 datasets were used to train and validate this system, containing 2965 samples from lung adenocarcinoma, skin cutaneous melanoma and head and neck squamous cell carcinoma. RESULTS: The patients with high immune and low stromal scores (HL) were associated with low ratio of T cell co-inhibitory/stimulatory molecules and low levels of angiogenesis markers, while the patients with low immune and high stromal scores (LH) had the opposite characteristics. The HL patients had immune-centered networks, while the patients with low immune and low stromal scores (LL) had desert-like networks. Moreover, copy number alteration burden was decreased in the HL patients. For the clinical characteristics, our TME classification was an independent prognostic factor. In the 5 cohorts with immunotherapy, the LH patients were linked to the lowest response rate. CONCLUSIONS: ISTMEscore system could reflect the TME status and predict the prognosis. Compared to previous TME scores, our ISTMEscore was superior in the prediction of prognosis and immunotherapy response.

Inhibition of miR-21 alleviated cardiac perivascular fibrosis via repressing EndMT in T1DM.

In type 1 and type 2 diabetes mellitus, increased cardiac fibrosis, stiffness and associated diastolic dysfunction may be the earliest pathological phenomena in diabetic cardiomyopathy. Endothelial-mesenchymal transition (EndMT) in endothelia cells (ECs) is a critical cellular phenomenon that increases cardiac fibroblasts (CFs) and cardiac fibrosis in diabetic hearts. The purpose of this paper is to explore the molecular mechanism of miR-21 regulating EndMT and cardiac perivascular fibrosis in diabetic cardiomyopathy. In vivo, hyperglycaemia up-regulated the mRNA level of miR-21, aggravated cardiac dysfunction and collagen deposition. The condition was recovered by inhibition of miR-21 following with improving cardiac function and decreasing collagen deposition. miR-21 inhibition decreased cardiac perivascular fibrosis by suppressing EndMT and up-regulating SMAD7 whereas activating p-SMAD2 and p-SMAD3. In vitro, high glucose (HG) up-regulated miR-21 and induced EndMT in ECs, which was decreased by inhibition of miR-21. A highly conserved binding site of NF-κB located in miR-21 5'-UTR was identified. In ECs, SMAD7 is directly regulated by miR-21. In conclusion, the pathway of NF-κB/miR-21/SMAD7 regulated the process of EndMT in T1DM, in diabetic cardiomyopathy, which may be regarded as a potential clinical therapeutic target for cardiac perivascular fibrosis.

Time-to-first appropriate shock in patients implanted prophylactically with an implantable cardioverter-defibrillator: data from the Survey on Arrhythmic Events in BRUgada Syndrome (SABRUS).

AIMS: Data on predictors of time-to-first appropriate implantable cardioverter-defibrillator (ICD) therapy in patients with Brugada Syndrome (BrS) and prophylactically implanted ICD's are scarce. METHODS AND RESULTS: SABRUS (Survey on Arrhythmic Events in BRUgada Syndrome) is an international survey on 678 BrS patients who experienced arrhythmic event (AE) including 252 patients in whom AE occurred after prophylactic ICD implantation. Analysis was performed on time-to-first appropriate ICD discharge regarding patients' characteristics. Multivariate logistic regression models were utilized to identify which parameters predicted time to arrhythmia ≤5 years. The median time-to-first appropriate ICD therapy was 24.8 ± 2.8 months. A shorter time was observed in patients from Asian ethnicity (P < 0.05), those with syncope (P = 0.001), and those with Class IIa indication for ICD (P = 0.001). A longer time was associated with a positive family history of sudden cardiac death (P < 0.05). Multivariate Cox regression revealed shorter time-to-ICD therapy in patients with syncope [odds ratio (OR) 1.65, P = 0.001]. In 193 patients (76.6%), therapy was delivered during the first 5 years. Factors associated with this time were syncope (OR 0.36, P = 0.001), spontaneous Type 1 Brugada electrocardiogram (ECG) (OR 0.5, P < 0.05), and Class IIa indication (OR 0.38, P < 0.01) as opposed to Class IIb (OR 2.41, P < 0.01). A near-significant trend for female gender was also noted (OR 0.13, P = 0.052). Two score models for prediction of <5 years to shock were built. CONCLUSION: First appropriate therapy in BrS patients with prophylactic ICD's occurred during the first 5 years in 76.6% of patients. Syncope and spontaneous Type 1 Brugada ECG correlated with a shorter time to ICD therapy.

αB-Crystallin Interacts with Nav1.5 and Regulates Ubiquitination and Internalization of Cell Surface Nav1.5.

Nav1.5, the pore-forming α subunit of the cardiac voltage-gated Na(+) channel complex, is required for the initiation and propagation of the cardiac action potential. Mutations in Nav1.5 cause cardiac arrhythmias and sudden death. The cardiac Na(+) channel functions as a protein complex; however, its complete components remain to be fully elucidated. A yeast two-hybrid screen identified a new candidate Nav1.5-interacting protein, αB-crystallin. GST pull-down, co-immunoprecipitation, and immunostaining analyses validated the interaction between Nav1.5 and αB-crystallin. Whole-cell patch clamping showed that overexpression of αB-crystallin significantly increased peak sodium current (INa) density, and the underlying molecular mechanism is the increased cell surface expression level of Nav1.5 via reduced internalization of cell surface Nav1.5 and ubiquitination of Nav1.5. Knock-out of αB-crystallin expression significantly decreased the cell surface expression level of Nav1.5. Co-immunoprecipitation analysis showed that αB-crystallin interacted with Nedd4-2; however, a catalytically inactive Nedd4-2-C801S mutant impaired the interaction and abolished the up-regulation of INa by αB-crystallin. Nav1.5 mutation V1980A at the interaction site for Nedd4-2 eliminated the effect of αB-crystallin on reduction of Nav1.5 ubiquitination and increases of INa density. Two disease-causing mutations in αB-crystallin, R109H and R151X (nonsense mutation), eliminated the effect of αB-crystallin on INa This study identifies αB-crystallin as a new binding partner for Nav1.5. αB-Crystallin interacts with Nav1.5 and increases INa by modulating the expression level and internalization of cell surface Nav1.5 and ubiquitination of Nav1.5, which requires the protein-protein interactions between αB-crystallin and Nav1.5 and between αB-crystallin and functionally active Nedd4-2.

Post-transcriptional regulation of cardiac sodium channel gene SCN5A expression and function by miR-192-5p.

The SCN5A gene encodes cardiac sodium channel Nav1.5 and causes lethal ventricular arrhythmias/sudden death and atrial fibrillation (AF) when mutated. MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression, and involved in the pathogenesis of many diseases. However, little is known about the regulation of SCN5A by miRNAs. Here we reveal a novel post-transcriptional regulatory mechanism for expression and function of SCN5A/Nav1.5 via miR-192-5p. Bioinformatic analysis revealed that the 3'-UTR of human and rhesus SCN5A, but not elephant, pig, rabbit, mouse, and rat SCN5A, contained a target binding site for miR-192-5p and dual luciferase reporter assays showed that the site was critical for down-regulation of human SCN5A. With Western blot assays and electrophysiological studies, we demonstrated that miR-192-5p significantly reduced expression of SCN5A and Nav1.5 as well as peak sodium current density INa generated by Nav1.5. Notably, in situ hybridization, immunohistochemistry and real-time qPCR analyses showed that miR-192-5p was up-regulated in tissue samples from AF patients, which was associated with down-regulation of SCN5A/Nav1.5. These results demonstrate an important post-transcriptional role of miR-192-5p in post-transcriptional regulation of Nav1.5, reveal a novel role of miR-192-5p in cardiac physiology and disease, and provide a new target for novel miRNA-based antiarrhythmic therapy for diseases with reduced INa.

Improved geometrical model of fringe projection profilometry.

The accuracy performance of fringe projection profilometry (FPP) depends on accurate phase-to-height (PTH) mapping and system calibration. The existing PTH mapping is derived based on the condition that the plane formed by axes of camera and projector is perpendicular to the reference plane, and measurement error occurs when the condition is not met. In this paper, a new geometric model for FPP is presented to lift the condition, resulting in a new PTH mapping relationship. The new model involves seven parameters, and a new system calibration method is proposed to determine their values. Experiments are conducted to verify the performance of the proposed technique, showing a noticeable improvement in the accuracy of 3D shape measurement.

Pravastatin inhibits plaque rupture and subsequent thrombus formation in atherosclerotic rabbits with hyperlipidemia.

Previous studies have demonstrated that statin can reduce the risk of acute coronary syndrome. In order to explore the mechanism, we observed the effects of pravastatin on plaque stability in atherosclerotic rabbits. Sixteen male rabbits were fed with a high fat diet following their damaged abdominal aortic endothelium by using catheter. Eight of them were administered with pravastatin (10 mg·kg(-1)·d(-1)) for 4 weeks. Then the rabbit atherosclerotic plaque rupture and thrombosis were triggered by injection of viper venom and histamine. Compared with model group, the thrombus area on aorta in pravastatin-treated group was reduced. Fibre cap on plaque was more thick and integrant, and inflammatory cell infiltration was also decreased. Serum total cholesterol, triglyceride, low density lipoprotein-cholesterol and contents of cholesterol in abdominal aorta were decreased. 6-Keto-prostaglandin F(1α) (6-keto-PGF(1α)) level and ratio of 6-keto-PGF(1α)/thromboxane B(2) (TXB(2)) in aorta were significantly increased. These results suggested that pravastatin could increase plaque stability and inhibit thrombosis through both lipid-dependent and lipid-independent way.

Effect of Xiaoaiping injection on advanced hepatocellular carcinoma in patients.

OBJECTIVE: To investigate the effect of Xiaoaiping Injection (XAP) on advanced hepatocellular carcinoma (HCC) in patients. METHODS: Sixty-eight patients with advanced HCC were assigned to a control group of 36 and a treatment group of 32. The control group was treated with best supportive treatment (BST) and the treatment group was given XAP plus BST. XAP was administered daily by i.v. and the treatment course was lasted for 30 days for both groups. The immediate therapeutic efficacy, Karnofsky performance status (KPS) scores, and the changes in immunity indicators (CD3+, CD4+ and CD8+ T cells) were measured and compared before and after treatment. The progression-free survival (PFS) rate and overall survival (OS) rate in the 2 groups were analyzed. RESULTS: The immediate therapeutic efficacy and KPS of the treatment group were better than those in the control (P < 0.05). Patients in the treatment group had higher percentages of CD3 and CD4 T-lymphocytes in peripheral blood than those in the control group (P < 0.05). The median survival time was 27.0 weeks in the treatment group and 24.5 weeks in the control group. The 6-months cumulative survival rates in the treatment and control groups were 33.3% and 25.0%, respectively, with no significant difference (P > 0.05). The PFS was 18 weeks in the treatment group and 15 weeks in control group (P < 0.05). CONCLUSION: XAP enhances the quality of life (QOL) of patients with advanced HCC, improves their immunity and extends their PFS.

[The effect of tanshinone II A on the transmembrance action potential of myocardial cells in LQT2 rabbit models].

OBJECTIVE: To determine the effect of tanshinone II A on the transmembrance action potential of three layers of myocardial cells in LQT2 rabbit models. METHODS: Whole-cell patch clamp technique was used to record the action potential duration 90 (APD90) and the transmural dispersion of repolarization (TDR) of the epicardium, mid-myocardium and endocardium in the LQT2 rabbits. RESULTS: Of the three layers of myocardial cells, the mid-myocardium had the longest APD90 and the epicardium had the shortest APD90. The APD90 of all three layers of myocardial cells were shortened with infusion of 10 mg/mL tanshinone II A. Their TDR values also reduced by the infusion. CONCLUSION: Tanshinone II A shortens APD90 and reduces TDR values.

A new risk score for the assessment of outcomes for Chinese patients with peripartum cardiomyopathy.

BACKGROUND: Peripartum cardiomyopathy (PPCM) is a potentially life-threatening complication of pregnancy, but identifying patients at higher risk of this condition remains difficult. OBJECTIVES: We conducted a study to identify new risk factors associated with PPCM and predictors of poor outcomes. METHODS: This retrospective analysis included a total of 44 women with PPCM. As a control group, 79 women who gave birth around the same time as the PPCM patients and who did not have organic disease were included. A multivariate regression analysis was conducted to identify risk factors associated with PPCM and with delayed recovery. RESULTS: All PPCM patients were discharged within 28 days. In comparison to the control group, PPCM patients had higher rates of preeclampsia (20.4% vs. 1.27%, P<0.001), autoimmune disease (27.3% vs. 11.4%, P = 0.018), and cesarean delivery with preterm labor (31.8% vs. 17.7%, P = 0.037). The neonates of PPCM patients had lower birth weight (2.70±0.66 kg vs. 3.21±0.57 kg, P<0.001). PPCM patients had higher levels of C-reactive protein, d-dimer, brain natriuretic peptide (BNP), and serum phosphorus, but lower levels of albumin and serum calcium (all P<0.001). In all patients with PPCM, the left ventricular ejection fraction (LVEF) returned to normal (≥50%) within 28 days after admission. Subjects with early recovery (n = 34) had lower BNP than those with delayed recovery (n = 10) (649.7 ± 526.0 pg/mL vs. 1444.1 ± 1040.8 pg/mL, P = 0.002). Multivariate regression led to a three-point score system to predict PPCM (1 point each for the presence of pericardial effusion, left ventricular dilatation, and d-dimer level ≥0.5 µg/mL). At a cutoff of ≥2, this scoring system predicted delayed recovery with 95.5% sensitivity and 96.1% specificity. The negative predictive value was 97.4% and the positive predictive value was 93.3%. Binary logistic regression indicated that PPCM patients with pulmonary hypertension, lower hemoglobin, or worse LVEF tended to require longer hospital stay (minimum 14 days). CONCLUSIONS: A risk score that consists of pericardial effusion, left ventricular dilatation, and d-dimer level ≥ 0.5 µg/mL could help streamline the diagnosis of PPCM prior to confirmatory investigations. Moreover, a risk score that consists of pulmonary hypertension, lower hemoglobin and worse LVEF could help to predict poor outcomes in PPCM patients.