Multi-Shell Nanourchin-Integrated Dual Mode Lateral Flow Immunoassay for Sensitive and Rapid Detection of Clinical Cardiac Myosin-Binding Protein C.

Cardiac myosin-binding protein C (cMyBP-C) is a novel cardiac marker of acute myocardial infarction (AMI) and acute cardiac injuries (ACI). Construction of point-of-care testing techniques capable of sensing cMyBP-C with high sensitivity and precision is urgently needed. Herein, we synthesized an Au@NGQDs@Au/Ag multi-shell nanoUrchins (MSNUs), and then applied it in a colorimetric/SERS dual-mode immunoassay for detection of cMyBP-C. The MSNUs displayed superior stability, colorimetric brightness, and SERS enhancement ability with an enhanced factor of 5.4 × 109, which were beneficial to improve the detection capability of test strips. The developed MSNU-based test strips can achieve an ultrasensitive immunochromatographic assay of cMyBP-C in both colorimetric and SERS modes with the limits of detection as low as 19.3 and 0.77 pg/mL, respectively. Strikingly, this strip was successfully applied to analyze actual plasma samples with significantly better sensitivity, negative predictive value, and accuracy than commercially available gold test strips. Notably, this method possessed a wide range of application scenarios via combining with a color recognizer application named Color Grab on the smartphone, which can meet various needs of different users. Overall, our MSNU-based test strip as a mobile health monitoring tool shows excellent sensitivity, reproducibility, and rapid detection of the cMyBP-C, which holds great potential for the early clinic diagnosis of AMI and ACI.

Machine learning in risk prediction of continuous renal replacement therapy after coronary artery bypass grafting surgery in patients.

OBJECTIVES: This study aimed to develop machine learning models for risk prediction of continuous renal replacement therapy (CRRT) following coronary artery bypass grafting (CABG) surgery in intensive care unit (ICU) patients. METHODS: We extracted CABG patients from the electronic medical record system of the hospital. The endpoint of this study was the requirement for CRRT after CABG surgery. The Boruta method was used for feature selection. Seven machine learning algorithms were developed to train models and validated using 10 fold cross-validation (CV). Model discrimination and calibration were estimated using the area under the receiver operating characteristic curve (AUC) and calibration plot, respectively. We used the SHapley Additive exPlanations (SHAP) method to illustrate the effects of the features attributed to the model and analyze the effects of individual features on the output of the mode. RESULTS: In this study, 72 (37.89%) patients underwent CRRT, with a higher mortality compared to those patients without CRRT. The Gaussian Naïve Bayes (GNB) model with the highest AUC were considered as the final predictive model and performed best in predicting postoperative CRRT. The analysis of importance revealed that cardiac troponin T, creatine kinase isoenzyme, albumin, low-density lipoprotein cholesterol, NYHA, serum creatinine, and age were the top seven features of the GNB model. The SHAP force analysis illustrated how created model visualized individualized prediction of CRRT. CONCLUSIONS: Machine learning models were developed to predict CRRT. This contributes to the identification of risk variables for CRRT following CABG surgery in ICU patients and enables the optimization of perioperative managements for patients.

High-fat diet causes mitochondrial damage and downregulation of mitofusin-2 and optic atrophy-1 in multiple organs.

High-fat consumption promotes the development of obesity, which is associated with various chronic illnesses. Mitochondria are the energy factories of eukaryotic cells, maintaining self-stability through a fine-tuned quality-control network. In the present study, we evaluated high-fat diet (HFD)-induced changes in mitochondrial ultrastructure and dynamics protein expression in multiple organs. C57BL/6J male mice were fed HFD or normal diet (ND) for 24 weeks. Compared with ND-fed mice, HFD-fed mice exhibited increased body weight, cardiomyocyte enlargement, pulmonary fibrosis, hepatic steatosis, renal and splenic structural abnormalities. The cellular apoptosis of the heart, liver, and kidney increased. Cellular lipid droplet deposition and mitochondrial deformations were observed. The proteins related to mitochondrial biogenesis (TFAM), fission (DRP1), autophagy (LC3 and LC3-II: LC3-I ratio), and mitophagy (PINK1) presented different changes in different organs. The mitochondrial fusion regulators mitofusin-2 (MFN2) and optic atrophy-1 (OPA1) were consistently downregulated in multiple organs, even the spleen. TOMM20 and ATP5A protein were enhanced in the heart, skeletal muscle, and spleen, and attenuated in the kidney. These results indicated that high-fat feeding caused pathological changes in multiple organs, accompanied by mitochondrial ultrastructural damage, and MFN2 and OPA1 downregulation. The mitochondrial fusion proteins may become promising targets and/or markers for treating metabolic disease.

Atorvastatin reverses high cholesterol-induced cardiac remodelling and regulates mitochondrial quality-control in a cholesterol-independent manner: An experimental study.

Mitochondria are key regulators of cell fate, maintaining self-stability by a fine-tuned quality-control network including mitophagy, biogenesis, fission and fusion processes. Myocardial mitochondria can be impaired by hypercholesterolemia. Statins, such as atorvastatin, are considered the cornerstone in the management of hypercholesterolaemia primarily due to their marked cholesterol-lowering ability. The direct effect of atorvastatin on myocardial mitochondria remains unclear. We aimed to explore whether atorvastatin could attenuate myocardial mitochondrial defects induced by high cholesterol, and whether cycloastragenol, a potent telomerase activator, could be used as a potential complementary bioactive compound for obesity and hypercholesterolaemia treatment. We found that atorvastatin at a low dose (3 mg/kg) did not reduce elevated serum cholesterol, but reversed cardiac remodelling and dysfunction in C57BL/6J mice fed with high-fat diet (HFD). Atorvastatin reversed the upregulated mitophagy, mitochondrial fission and fusion, accompanied by mitochondrial biogenesis activation in HFD-fed mice hearts. Mitochondrial structural impairments were attenuated by atorvastatin in HFD-fed mice and oxidized low-density lipoprotein (ox-LDL) exposed HL-1 cardiomyocytes. The depolarized mitochondrial membrane potential and increased mitochondrial oxygen consumption rates in ox-LDL exposed HL-1 cells were recovered by atorvastatin. Furthermore, atorvastatin co-treated with cycloastragenol had better effects on reducing body weight, improving cardiac remodelling and dysfunction, and protecting mitochondria in high cholesterol. Conclusively, low-dose atorvastatin exhibited a cholesterol-independent cardioprotective effect through improving the mitochondrial quality-control network and repairing mitochondrial ultrastructure in high cholesterol. Atorvastatin plus cycloastragenol supplement therapy has a better effect on treating obesity and hypercholesterolaemia.

Astragaloside IV Prevents Cardiac Remodeling in the Apolipoprotein E-Deficient Mice by Regulating Cardiac Homeostasis and Oxidative Stress.

BACKGROUND: Hypercholesterolemia is a risk factor for the development of cardiac hypertrophy. Astragaloside IV (AST-IV) possesses cardiovascular protective properties. We hypothesize that AST-IV prevents cardiac remodeling with hypercholesterolemia via modulating tissue homeostasis and alleviating oxidative stress. METHODS: The ApoE-/- mice were treated with AST-IV at 1 or 10 mg/kg for 8 weeks. The blood lipids tests, echocardiography, and TUNEL were performed. The mRNA expression profile was detected by real-time PCR. The myocytes size and number, and the expressions of proliferation (ki67), senescence (p16INK4a), oxidant (NADPH oxidase 4, NOX4) and antioxidant (superoxide dismutase, SOD) were observed by immunofluorescence staining. RESULTS: Neither 1 mg/kg nor 10 mg/kg AST-IV treatment could decrease blood lipids in ApoE-/- mice. However, the decreased left ventricular ejection fraction (LVEF) and fractional shortening (FS) in ApoE-/- mice were significantly improved after AST-IV treatment. The cardiac collagen volume fraction declined nearly in half after AST-IV treatment. The enlarged myocyte size was suppressed, and myocyte number was recovered, and the alterations of genes expressions linked to cell cycle, proliferation, senescence, p53-apoptosis pathway and oxidant-antioxidants in the hearts of ApoE-/- mice were reversed after AST-IV treatment. The decreased ki67 and increased p16INK4a in the hearts of ApoE-/- mice were recovered after AST-IV treatment. The percentages of apoptotic myocytes and NOX4-positive cells in AST-IV treated mice were decreased, which were consistent with the gene expressions. CONCLUSION: AST-IV treatment could prevent cardiac remodeling and recover the impaired ventricular function induced by hypercholesterolemia. The beneficial effect of AST-IV might partly be through regulating cardiac homeostasis and anti-oxidative stress.

Atorvastatin ameliorates lipid overload-induced mitochondrial dysfunction and myocardial hypertrophy by decreasing fatty acid oxidation through inactivation of the p-STAT3/CPT1 pathway.

Although statins are shown to have cardiac pleiotropic effects independent of lowering cholesterol, the underlying mechanism remains unclear. Mitochondrial dysfunction induced by increased fatty acid oxidation (FAO) is the culprit in the development of cardiac hypertrophy and dysfunction. This study was to explore whether the cardiac pleiotropic effects of atorvastatin were associated with FAO regulation, with a specific focus on carnitine palmitoyltransferase 1 (CPT1). High-fat diet (HFD)-fed mice and palmitic acid (PA)-stimulated neonatal rat primary cardiomyocytes (NRCMs) were treated with atorvastatin, with or without FAO modulators, signal transducer and activator of transcription 3 (STAT3) agonist, and inhibitor. Atorvastatin (3 mg/kg) did not reduce serum cholesterol levels in HFD-fed mice but ameliorated mitochondrial dysfunction and cardiac hypertrophy. In vitro, atorvastatin and the FAO inhibitor alleviated PA-induced mitochondrial dysfunction and cardiomyocyte hypertrophy. However, the FAO enhancer eliminated atorvastatin's protective effects. Furthermore, atorvastatin decreased CPT1 and FAO levels and prevented STAT3 phosphorylation and nuclear translocation. STAT3 inhibitor had the same inhibitory effects as atorvastatin on CPT1, FAO levels, and cardiomyocyte hypertrophy, whereas STAT3 agonist disrupted these effects of atorvastatin. Our results demonstrate that atorvastatin decreases myocardial FAO by inactivating the p-STAT3/CPT1 signaling pathway, which improves lipid overload-induced mitochondrial dysfunction and cardiac hypertrophy in a cholesterol-independent manner. This is the first study to explore the cardiac pleiotropic effects of atorvastatin with respect to FAO. However, whether atorvastatin regulates FAO in the cardiac hypertrophy model induced by other variables has not been investigated in this work, and this is expected to be performed in the future.

Low-dose atorvastatin protects skeletal muscle mitochondria in high-fat diet-fed mice with mitochondrial autophagy inhibition and fusion enhancement.

Despite the great clinical benefits of statins in cardiovascular diseases, their widespread use may lead to adverse muscle reactions associated with mitochondrial dysfunction. Some studies have demonstrated that statins provide substantial improvement to skeletal muscle health in mice. Our previous study found that oral treatment with atorvastatin (Ator, 3 mg/kg) protected myocardial mitochondria in high-fat diet (HFD)-fed mice. Therefore, this study aimed to explore the influence of low-dose Ator (3 mg/kg) on mitochondria in skeletal muscle under cholesterol overload. Male C57BL/6J mice were fed a HFD for 18 weeks and orally administered Ator (3 mg/kg) during the last 12 weeks. Ator treatment had no effects on elevated serum cholesterol and glucose levels in HFD-fed mice. Serum creatine kinase levels and the cross-sectional area of muscle cells were not affected by HFD feeding or Ator treatment. Increased expression of PINK1-LC3 II (activated mitophagy), MFN2 (fusion), and PGC-1α (biogenesis) proteins was induced in the skeletal muscles of HFD-fed mice. Treatment with Ator inhibited PINK1 and LC3 II protein expression, but further promoted MFN1, MFN2, and OPA1 expression. The impairments in mitochondrial quality and morphology in HFD-fed mice were attenuated by treatment with Ator. Furthermore, Ator treatment enhanced glucose oxidation capacity and restored ATP production in the skeletal muscles of HFD-fed mice. The study reveals that low-dose Ator has a protective effect on muscle mitochondria in mice, likely through inhibiting mitophagy and enhancing mitochondrial fusion. This suggests that skeletal muscle mitochondria may be one of low-dose Ator-mediated protective targets.

Astragaloside IV alleviates hypoxia/reoxygenation-induced neonatal rat cardiomyocyte injury via the protein kinase a pathway.

BACKGROUND: Astragaloside IV (As-IV) exerts beneficial effects on hypoxia/reoxygenation (H/R)-induced cardiomyocyte injury, possibly through normalization of sarcoplasmic reticulum Ca(2+) ATPase (SERCA2a) function. The exact mechanisms remain unknown. This study was designed to investigate the role of protein kinase A (PKA) in the protective effect of As-IV on SERCA2a function. METHODS: Cultured cardiomyocytes from neonatal rats were exposed to 6 h of hypoxia followed by 3 h of reoxygenation (H/R) with or without As-IV treatment. Myocyte injury was determined by the creatine kinase (CK)-MB fraction in supernatant. Myocardial SERCA2a activity and PKA kinase activity were assessed. PKA subunit mRNA expression and Ser(16) phosphorylated phospholamban (Ser(16)-PLN) protein expression were detected by real-time PCR and Western blot, respectively. RESULTS: The administration of As-IV significantly decreased CK-MB release and restored SERCA2a activity in H/R cardiomyocytes. The mRNAs of PKA subunits, PKA-RIα, PKA-RIIα, PKA-RIIß, PKA-Cα and PKA-Cß, were downregulated in H/R cardiomyocytes. However, PKA-Cα mRNA expression was significantly increased after As-IV treatment. Meanwhile, there was a tendency to recovery of the H/R-induced PKA kinase activity decrease after As-IV treatment. The expression of Ser(16)-PLN protein, which is specifically phosphorylated by PKA, was upregulated in As-IV-treated H/R cardiomyocytes. CONCLUSIONS: These results suggest that the cardioprotection of As-IV may be through the upregulation of PKA and Ser(16)-PLN, thereby restoring SERCA2a function in H/R injury.

Analysis of gene expression profiling of amyloidogenic immunoglobulin light-chains on cultured rat cardiomyocytes.

The present study aimed to investigate the toxic effects of different amyloidogenic light-chains (LCs) on cardiomyocytes, and demonstrate the differentially expressed genes (DEGs) and signaling pathways that participate in this process. Cultured cardiomyocytes were treated with recombinant κ LC peptide (AL-09) or with serum from a patient diagnosed with multiple myeloma (λ LC) with cardiac involvement. The 6xHis peptide or serum from healthy patients was used as peptide control or serum control, respectively. Cell viability was determined using CCK-8 assay and apoptosis was analyzed by flow cytometry. The DEGs were detected by RNA sequencing (RNA-Seq), followed by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Changes in gene expression levels were confirmed by reverse transcription-quantitative PCR. The cell viability in the AL-09 peptide-treated (0.2 mg/ml) and patient serum-treated (1:10 dilution) cardiomyocytes decreased to 42 and -72% of the corresponding control groups. The extent of cell apoptosis increased in AL-09-treated cardiomyocytes compared with the control group. RNA-Seq showed 256 DEGs co-existed in the two paired groups, including 127 upregulated and 88 downregulated genes. The KEGG pathways for upregulated expressed genes included the 'TGF-ß signaling pathway', the 'Hedgehog signaling pathway', the 'ErbB signaling pathway' and 'lysine degradation'. The higher mRNA expression of bone morphogenetic protein (Bmp) 4, Bmp6, prostaglandin G/H synthase (Ptgs)1, Ptgs2, epiregulin, Tgfa and procollagen-lysine,2-oxoglutarate 5-dioxygenase 2 were confirmed. The KEGG pathways of downregulated expressed genes included genes involved with the 'p53 signaling pathway' and the 'cell cycle'. The mRNA expression levels of E3 ubiquitin-protein ligase CCNB1IP1 showed significant downregulation in the AL-09 peptide group compared with those in the 6xHis peptide group. In conclusion, cardiomyocytes treated with amyloidogenic λ and κ LCs presented with decreased cell viability compared with controls. Cell apoptosis increased in κ LC-treated cells compared with controls. The gene expression profiles associated with transforming growth factor-ß-bone morphogenetic protein, the receptor tyrosine-protein kinase erbB-2 signaling pathways, prostaglandins, collagen production, the p53 signaling pathway and the cell cycle were altered in light-chain-treated cardiomyocytes.

Cardiac Myosin-Binding Protein C Release Profile After Cardiac Surgery in Intensive Care Unit.

BACKGROUND: Cardiac surgical procedures produce iatrogenic myocardial cell injury with necrosis that result in an obligatory release of biomarkers. Cardiac myosin binding protein C (cMyBP-C) has recently emerged as a specific and sensitive biomarker in patients with acute myocardial injury. We therefore aimed to investigate the release profiles of cMyBP-C after cardiac surgical procedures. METHODS: Enzyme-linked immunosorbent assay to detect blood cMyBP-C was established by using two monoclonal antibodies against N-terminus of human cMyBP-C. Consecutive patients undergoing cardiac operations (N = 151) were recruited in this study. Blood cMyBP-C was assayed preoperatively, at intensive care unit arrival (0 hour after the operation), at 2 to 48 hours, and before discharge. The characteristics and detailed surgical procedure were recorded. RESULTS: The established immunoassay was capable of detecting human cMyBP-C (0 to 1000 ng/L). The released cMyBP-C peaked immediately after cardiac surgery (0 h), attaining 3.8-fold higher than before the operation, dropped abruptly within 24 hours, and stayed at a higher level until discharge. Postoperative cMyBP-C levels correlated positively with high-sensitivity cardiac troponin T (hs-cTnT), creatine kinase, myoglobin, and creatine kinase MB isoenzyme. Different cardiac surgical procedures were characterized by different levels of release of cardiac biomarkers. Isolated off-pump coronary artery bypass grafting was associated with the smaller amount of cMyBP-C release, whereas valve replacement/plasty surgery produced higher release, in particular the multiple-valve surgery. Both cMyBP-C and hs-cTnT correlated with surgical techniques, postoperative intensive care unit stay, and hospital stay. CONCLUSIONS: Circulating cMyBP-C is a promising novel biomarker for evaluating cardiac surgical trauma in patients undergoing a cardiac operation.

[Reduced Ca2+ current in rat cardiomyocytes transfected with troponin I R145W mutation gene].

OBJECTIVE: To investigate the effects of cardiac troponin I R145W mutation, detected in Chinese patients with hypertrophic cardiomyopathy, on Ca(2+) current modulation. METHODS: R146W mutation (resemble R145W in human) was introduced into rat cardiac troponin I cDNA by site-directed mutagenesis. With EGFP as a reporter gene, replication-defective adenovirus containing the wild or mutant cTnI gene was constructed. Adult rat cardiomyocytes, were isolated by Langendorff perfusion and cultured with serum-free medium and transduced with the recombinant adenoviruses. Western blot was used to determine the recombinant proteins. Whole cell patch clamp was employed to record L-type Ca(2+) currents on cultured myocytes. Intracellular free Ca(2+) and caffeine-induced sarcoplasmic reticulum (SR) Ca(2+) release were determined after the cells incubated with Fura-2/AM. RESULTS: DNA sequencing confirmed that R146W mutation was generated in rat cTnI cDNA. Bright green fluorescence was observed in the cultured cardiomyocytes at 48 h after transduction. The recombinant proteins could be identified with cTnI or GFP monoclonal antibody. The peak current of L-type Ca(2+) channel in cells transduced with cTnI R146W was significantly decreased compared to control cells and cells transfected with wild cTnI. Intracellular free Ca(2+) concentrations and caffeine-induced SR Ca(2+) release determined by Fura-2/AM were similar among various cells. CONCLUSION: Reduced peak current of L-type Ca(2+) channel in cells transduced with cTnI R146W might contribute to the disease-causing mechanism of this mutation in patients with hypertrophic cardiomyopathy.

[Association between cardiac troponin I phosphorylation/degradation and cardiomyopathies].

OBJECTIVE: To investigate the association between cTnI phosphorylation/degradation and cardiomyopathies in extransplanted myocardium. METHODS: cTnI phosphorylation and degradation as well as PKC (beta1, beta2) expressions were determined in extransplanted hearts from patients with cardiomyopathies (n = 8) and from traffic accidents (n = 6) by Western blot. RESULTS: The cTnI bands were observed in LV myocardium of cardiomyopathy patients and normal myocardium while and cTnI degradation bands were only detected in LV myocardium from patients with cardiomyopathies. The phosphorylated cTnI bands were significantly upregulated in LV myocardium of cardiomyopathy patients compared to normal myocardium (P < 0.05). There was no myocardial PKCbeta1, PKCbeta2 expression in all examined hearts. CONCLUSION: The cTnI degradation products and increased phosphorylated cTnI expression are likely involved in the pathogenesis and development of cardiomyopathy.

Preparation of Monoclonal Antibodies and a Simple Myeloperoxidase-Immunosorbent Assay for Detecting Human Myeloperoxidase.

Myeloperoxidase (MPO), a leukocyte hemoprotein released from neutrophils, is thought to be a potential participant in plaque formation and plaque rupture. Therefore, MPO is regarded as an early marker predicting the risk for atherosclerosis, especially for coronary artery disease and acute coronary syndrome. We generated hybridoma clones 1E3 and 3E8 secreting monoclonal antibodies (mAbs) specific to human MPO. BALB/c mice were immunized with MPO protein purified from human neutrophils. Splenocytes from these mice were fused with the mouse myeloma cell line SP2/0. Based on isotyping of the mAbs, both clones 1E3 and 3E8 were referred to the IgG1 subclass. The specificities of 1E3 and 3E8 were assessed by enzyme-linked immunosorbent assay (ELISA), and only 3E8 was confirmed by western blot. We developed a simple MPO-immunosorbent assay (MPO-ISA) on microplate based on both the immune activity and peroxidase activity of MPO. The mAb secreted by clone 3E8 was chosen as coating antibody to capture the plasma MPO without interfering with the peroxidase activity of MPO. Then, tetramethylbenzidine substrate was added to the microwell directly, catalyzed by captured MPO, and a colored product was formed. The simple MPO-ISA test has a sensitivity of 3.68 ng/mL. The linear concentration of MPO-ISA for commercial MPO standard ranged to 250 ng/mL. The average recovery rate is 101.02%. The imprecision within-day was <10% at three different MPO levels. The imprecision between-day was <10% at low and middle MPO levels and varied to 14.61% at the high MPO level. We found that the established MPO-ISA can detect the plasma MPO from human and cavy, but not from mouse and rat. Compared with the commercial human MPO ELISA assay, the MPO-ISA can be used to detect the natural human MPO protein, but not recombinant MPO polypeptides. The generated mAbs and MPO-ISA test may be useful tools to assess risk for inflammation and cardiac events.

The effects of different initiation time of exercise training on left ventricular remodeling and cardiopulmonary rehabilitation in patients with left ventricular dysfunction after myocardial infarction.

PURPOSE: The purpose of this study was to determine whether different initiation of exercise training (ET) produces different effect sizes for left ventricular (LV) remodeling and cardiopulmonary rehabilitation in patients with LV dysfunction after myocardial infarction (MI). METHOD: Trials evaluating ET outcomes identified by searches in OVID MEDLINE, EMBASE, PubMed and WEB OF SCIENCE were used. Meta-analysis was conducted with the use of the software STATA 11.0. The results were expressed as the standardized mean difference (SMD), with corresponding 95% CI and p value. RESULTS: The largest changes in LV remodeling and cardiopulmonary capacity rehabilitation were obtained when programs began the acute phase after MI. With the healing of MI, the beneficial effects of ET on LV ejection fraction (LVEF), LV end-systolic diameter (LVDs) and peak VO2 were gradually weakened even worse. The incidence of major adverse cardiac events was not significantly increased in acute phase post-MI. Sensitivity analyses show that ET still had significant effect in reducing LVDs and increasing peak VO2, while ET no longer had statistical effect in increasing LVEF but showed favorable trends when the same research institution's works were excluded. CONCLUSIONS: ET has favorable effects on LV remodeling and cardiopulmonary rehabilitation in LV dysfunction post-MI patients. The greatest benefits are obtained when ET starts at the acute phase following MI. IMPLICATIONS FOR REHABILITATION: Early exercise training is safe and feasible in acute and healing phase after myocardial infarction. Early exercise training could attenuate LV remodeling and improve cardiopulmonary capacity in patients with myocardial infarction after hospital discharge (around one week post-MI). Exercise training has favorable effects on LV remodeling and cardiopulmonary capacity rehabilitation. Exercise training should be treated to have the same roles with drugs in secondary prevention of myocardial infarction.

In Vitro Effects of Hollow Gold Nanoshells on Human Aortic Endothelial Cells.

Gold nanoparticles are emerging as promising biomedical tools due to their unique nanoscale characteristics. Our purpose was to synthesize a hollow-shaped gold nanoparticle and to investigate its effect on human aortic endothelial cells (HAECs) in vitro. Hollow gold nanoshells with average 35-nm diameters and 10-nm shell thickness were obtained by galvanic replacement using quasi-spherical nanosilver as sacrifice-template. Our results showed that hollow gold nanoshells in the culture medium could be internalized into the cytoplasm of HAECs. No cytotoxicity effect of hollow gold nanoshells on HAECs was observed within the test concentrations (0-0.8 µg/mL) and test exposure period (0-72 h) by tetrazolium dye assay. Meanwhile, the release of cell injury biomarker, lactate dehydrogenase, was not significantly higher than that from control cells (without hollow gold nanoshells). The concentrations of vasodilators, nitric oxide, and prostacyclin I-2 were not changed, but the vasoconstrictor endothelin-1 was decreased by hollow gold nanoshells treatment in HAECs. HAECs exposed to hollow gold nanoshells resulted in suppressing expressions of genes involved in apoptosis and activating expressions of genes of adhesion molecules. Moreover, we demonstrated by in vitro endothelial tube formation that hollow gold nanoshells (0.8 µg/mL) could not inhibit angiogenesis by the HAECs. Altogether, these results indicate that the structure and major function of HAECs would not be disrupted by hollow gold nanoshell treatment.

Beneficial effects of astragaloside IV against angiotensin II-induced mitochondrial dysfunction in rat vascular smooth muscle cells.

Angiotensin II (Ang II)-induced mitochondrial dysfunction is a prominent characteristic of the majority of cardiovascular diseases. Astragaloside IV (As-IV), the major active ingredient of Astragalus membranaceus (Fisch.) Bge. (a traditional Chinese herbal medicine), possesses antioxidant properties. The present study was carried out to examine whether As-IV can reverse Ang II-induced mitochondrial dysfunction in vascular smooth muscle cells (VSMCs) and to elucidate the underlying molecular mechanisms. Cultured rat aortic VSMCs treated with Ang II (1 µM) for 24 h exhibited mitochondrial dysfunction, including a decrease in mitochondrial oxygen consumption rates (OCRs), adenosine triphosphate (ATP) production and mitochondrial DNA (mtDNA) levels, as well as the disruption of mitochondrial structural integrity. Following treatment with Ang II, As-IV (50 µg/ml) was added to the culture medium followed by incubation for a further 24 h. The administration of As-IV significantly increased the mitochondrial OCRs, ATP production and the mtDNA levels, and reversed the mitochondrial morphological changes which occurred in the VSMCs. Treatment with As-IV also reversed the Ang II-induced increase in the production of reactive oxygen species (ROS), the increase in NADPH oxidase and xanthine oxidase activity, as well as the decrease in mitochondrial membrane potential (ΔΨm) and manganese superoxide dismutase (Mn-SOD) activity. Furthermore, treatment with As-IV led to an increase in the mRNA expression of peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α) and mitochondrial transcription factor A (Tfam), and in the protein expression of PGC-1α, parkin and dynamin 1-like protein 1 (Drp1) in the VSMCs. These results indicate that As-IV exerts beneficial effects on Ang II-induced mitochondrial dysfunction in rat VSMCs and that these effects are mediated through the inhibition of ROS overproduction, as well as the promotion of mitochondrial autophagy and mitochondrial biogenesis. These data demonstrate the antioxidant properties of As-IV.

Oxygen-derived free radicals mediate endothelium-dependent contractions to acetylcholine in aortas from spontaneously hypertensive rats.

Experiments were designed to investigate whether or not oxygen-derived free radicals mediate endothelium-dependent contractions to acetylcholine in the aorta of spontaneously hypertensive rat (SHR). Isometric tension was measured in aortic rings taken from adult male SHR and Wistar-Kyoto rat (WKY) in the presence of NG-nitro-L-arginine. Endothelium-dependent contractions to acetylcholine were significantly greater in rings from SHR compared to WKY. Oxygen-derived free radicals, generated from xanthine plus xanthine oxidase, induced contractions that were larger in aortas from SHR than from WKY. Contractions to acetylcholine and free radicals were abolished by a selective TP-receptor antagonist, S 18886, and a preferential inhibitor of cyclo-oxygenase-1, valeryl salicylate, but not by a preferential inhibitor of cyclo-oxygenase-2, NS-398. Allopurinol, deferoxamine and the combination of superoxide dismutase plus catalase inhibited the contractions to oxygen-derived free radicals but did not significantly affect those to acetylcholine. In contrast, diethyldithiocarbamic acid, an inhibitor of superoxide dismutase, or Tiron, a scavenger of superoxide anion, reduced endothelium-dependent contractions to acetylcholine in aortas from SHR. The effect of these two drugs was additive. In SHR chronically treated with dimethylthiourea endothelium-dependent contractions to acetylcholine were decreased, and reduced further by acute in vitro exposure to deferoxamine or the combination of superoxide dismutase plus catalase. These results suggest that in the SHR aorta acetylcholine-induced endothelium-dependent contractions involve endothelial superoxide anion production and the subsequent dismutation into hydroxyl radicals and/or hydrogen peroxide. The free radicals activate cyclo-oxygenase-1, most likely to produce endoperoxides. Activation of TP-receptors is required to observe endothelium-dependent contractions to acetylcholine or endothelium-independent contractions in response to free radical generation.

Targeted efficacy of dihydroartemisinin for translationally controlled protein expression in a lung cancer model.

OBJECTIVE: Lung cancer is one of the malignant tumors with greatest morbidity and mortality around the world. The keys to targeted therapy are discovery of lung cancer biomarkers to facilitate improvement of survival and quality of life for the patients with lung cancer. Translationally controlled tumor protein (TCTP) is one of the most overexpressed proteins in human lung cancer cells by comparison to the normal cells, suggesting that it might be a good biomarker for lung cancer. MATERIALS AND METHODS: In the present study, the targeted efficacy of dihydroartemisinin (DHA) on TCTP expression in the A549 lung cancer cell model was explored. RESULTS AND CONCLUSIONS: DHA could inhibit A549 lung cancer cell proliferation, and simultaneously up-regulate the expression of TCTP mRNA, but down-regulate its protein expression in A549 cells. In addition, it promoted TCTP protein secretion. Therefore, TCTP might be used as a potential biomarker and therapeutic target for non-small cell lung cancers.

Periostin expression is upregulated and associated with myocardial fibrosis in human failing hearts.

BACKGROUND AND PURPOSE: Periostin, a matricellular protein, plays an important role in cardiac development and remodeling. Its expression profile and the association with myocardial fibrosis have not been investigated in human failing hearts. This work aimed to explore the behavior and pathologic significance of periostin in signifying collagen fibrogenesis in human hearts from patients with end-stage heart failure. METHODS AND SUBJECTS: Tissues were collected from heart transplant recipients and the control hearts were from unmatched donors. Periostin mRNA and protein were detected using quantitative real-time polymerase chain reaction and Western blotting. Immunohistochemistry staining was employed to directly detect the protein level and distribution of periostin in heart tissues. The extent of myocardial fibrosis was expressed by the percentage of Masson's trichrome staining. Gelatin zymography was used to detect the activities of matrix metalloproteinase (MMP)2 and MMP9. RESULTS: A low level of periostin mRNA expression was found in control hearts while not detectable at the protein level. Periostin mRNA was increased significantly in failing myocardium compared to that of controls. Periostin was distributed extensively in left ventricle and interventricular septum of the failing hearts. Correlation analysis showed periostin protein expression was positively associated with myocardial fibrosis as well as left ventricular diastolic dimension. The distribution and extent of periostin was consistent with that of myocardial fibrosis. MMP2 activity has an obvious increase about fourfold in heart tissues from HF patients. But there is no quantitative association with the expression of periostin. CONCLUSIONS: Periostin, the distribution and expression of which were consistent with the extent of myocardial fibrosis, might be a potential biomarker of cardiac remodeling in heart failure patients.

Cardiac troponin I is abnormally expressed in non-small cell lung cancer tissues and human cancer cells.

Cardiac troponin I (cTnI) is the only sarcomeric protein identified to date that is expressed exclusively in cardiac muscle. Its expression in cancer tissues has not been reported. Herein, we examined cTnI expression in non-small cell lung cancer (NSCLC) tissues, human adenocarcinoma cells SPCA-1 (lung) and BGC 823 (gastric) by immunohistochemistry, western blot analysis and real-time PCR. Immunopositivity for cTnI was demonstrated in 69.4% (34/49) NSCLC tissues evaluated, and was strong intensity in 35.3% (6/17) lung squamous cell carcinoma cases. The non-cancer-bearing lung tissues except tuberculosis (9/9, 100%) showed negative staining for cTnI. Seven monoclonal antibodies (mAbs) against human cTnI were applied in immunofluorescence. The result showed that the staining pattern within SPCA-1 and BGC 823 was dependent on the epitope of the cTnI mAbs. The membrane and nucleus of cancer cells were stained by mAbs against N-terminal peptides of cTnI, and cytoplasm was stained by mAbs against the middle and C-terminal peptides of cTnI. A ~25 kD band was identified by anti-cTnI mAb in SPCA-1 and BGC 823 extracts by western blot, as well as in cardiomyocyte extracts. The cTnI mRNA expressions in SPCA-1 and BGC 823 cells were about ten thousand times less than that in cardiomyocytes. Our study shows for the first time that cTnI protein and mRNA were abnormally expressed in NSCLC tissues, SPCA-1 and BGC 823 cells. These findings challenge the conventional view of cTnI as a cardiac-specific protein, enabling the potential use of cTnI as a diagnostic marker or targeted therapy for cancer.