ECV304 Cells Research Articles

Effects of Vaccinium myrtillus L. extract on TNF-α-induced inflammation human paraoxonases

Cardiovascular diseases (CVDs) is a leading cause of death worldwide, and is characterized by a range of conditions that affect the heart and blood vessels. The extract of Vaccinium myrtillus L. fruit has been found to have cardioprotective effect. Human paraoxonases (PON1, PON2, PON3) genes play an important role in protecting against the development of CVDs by acting as antioxidants and anti-inflammatory agents.The present study aimed to investigate the chemical constituents and cardioprotective effects of V. myrtillus L. extract on TNF-α inflammation and oxidative stress in ECV-304 human endothelial cell line and the potential role of human paraoxonases in these effects. The results indicated that V. myrtillus L. extract has cardioprotective effects against inflammation and oxidative stress in ECV-304 cells, and these effects may be mediated through the upregulation of PON1, PON2, and PON3 genes. These findings suggest that V.myrtillus L. may have therapeutic potential for the prevention and treatment of CVDs.

Endometriyal Hücrelerde Spirulina platensis Ekstresinin Kimyasal Bileşiminin Seçici Sitotoksisitesi: İn Vitro ve İn Siliko Yaklaşım

Microalgae are considered as an ample source of bioactive compounds. These wide variety of chemicals possess promising potential to translate into drug candidates. Hence, the aim of this study is to determine potential anti-cancer effects of Spirulina platensis microalgae extract against endometrial cancer cells. The selective cytotoxic effect and cytotoxicity index of S. platensis microalgae extract on the Ishikawa endometrial cancer cell line, the determination of DNA fragmentation and the change in gene expression levels of apoptosis-related Bad, Poly-ADP Ribose Polymerase-1 (PARP1), p53, Akt1 and caspase-3 signaling proteins were determined. ECV304 human umbilical vein endothelial cell line was used for a control cell line. Cytotoxic effects of 500 μg mL-1 of S. platensis microalgae extract on Ishikawa and ECV304 cells were determined as 43±4.87 % and 22±1.87 % respectively. In order to delineate specific compounds in the extract, GC-MS analysis was further conducted and seven major compounds revealed to be abundantly present. Since DNA fragmentation induced by S. platensis extract might be related interaction of GC-MS identified compounds with PARP1, an in silico analysis was further implemented. Results indicated the presence of a possible PARP1 inhibitory mechanism contributing to the apoptotic response. In conclusion, a possible link between a specific compound found in S. platensis microalgae extract and the cytotoxicity in endometrial cells was formed through combining in vitro and in silico approaches. The result of these approaches has contributed to the identification of potential anti-cancer compound candidates found in S. platensis microalgae extract.

An Online CMC–LC/MS Method for Screening Angiogenesis Promoting Components in Yiqi Fumai Lyophilization Injection (YQFM)

Objective In this study, an online CMC–LC/MS method was established in order to screen the active components of Yiqi Fumai lyophilization injection (YQFM) related to angiogenesis. The pro-angiogenic active components of YQFM were screened using this system and were then further verified. Methods The ECV304/CMC model coupled online with an HPLC–MS system was initially developed to screen, analyze, and identify the allergenic components of YQFM. A fraction was retained on the ECV304/CMC column, which was identified as ginsenoside Rb2. A cell proliferation assay and zebrafish intestinal microvascular model labeled with green fluorescence were used to detect the proliferation effect of ginsenoside Rb2. Results (1) The active component in YQFM, which was retained on the column, was ginsenoside Rb2. (2) Within a concentration range of 2.5–20 μg/mL, ginsenoside Rb2 was shown to promote the proliferation of ECV304 and EPCs cells and increase the area of sub-intestinal vessels in zebrafish. Conclusion Ginsenoside Rb2, the active component in YQFM, can promote angiogenesis. The ECV304/CMC-online-HPLC system may serve as an effective screening tool for the vasoactive components from the traditional Chinese medicine injection.

Lycopene alleviates oxidative stress-induced cell injury in human vascular endothelial cells by encouraging the SIRT1/Nrf2/HO-1 pathway

ABSTRACT Background and objective: Epidemiological research have displayed that dietary intake rich in lycopene, an antioxidant, is negatively correlated with the risk of cardiovascular disease (CVD). This study aimed to investigate whether the intervention with different concentrations of lycopene could attenuate H2O2-induced oxidative stress injury in human vascular endothelial cells (VECs). Methods: The human VECs HMEC-1 and ECV-304 were incubated with a final concentration of 300 µmol/L H2O2, followed by they were incubated with lycopene at doses of 0.5, 1, or 2 µm. Subsequently, cell proliferation, cytotoxicity, cell adhesion, reactive oxygen species (ROS) contents, adhesion molecule expression, oxidative stress levels, pro-inflammatory factor production, the apoptosis protein levels, and the silent information regulator-1 (SIRT1)/nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) pathway protein levels were tested by CCK-8 kit, lactate dehydrogenase (LDH) kit, immunofluorescence labeling, cell surface enzyme immunoassays (EIA), enzyme-linked immunosorbent assay (ELISA), as well as Western blot assays, respectively. Results: Under H2O2 stimulation, HMEC-1 and ECV-304 cell proliferation and the SIRT1/Nrf2/HO-1 pathway protein expression were significantly reduced, whereas cytotoxicity, apoptosis, cell adhesion molecule expression, pro-inflammatory and oxidative stress factors production were apparently encouraged, which were partially countered by lycopene intervention in a dose-dependent manner. Conclusion: Lycopene alleviates H2O2-induced oxidative damage in human VECs by reducing intracellular ROS levels, inflammatory factor production, cell adhesiveness, and apoptosis rate under oxidative stress conditions through activation of the SIRT1/Nrf2/HO-1 pathway.

Efficacy of Sodium Alginate Microsphere Interventional Embolization Combined with Sorafenib in Advanced Hepatocellular Carcinoma

To explore the efficacy of sodium alginate microsphere (SAM) transcatheter arterial chemoembolization (TACE) combined with sorafenib in liver cancer (LC), 118 patients with advanced LC were enrolled in the study. Patients were randomized into c-TACE and SAM-TACE groups; in addition to TACE, the SAM-TACE group was treated with SAM and oral sorafenib. The physical properties and cytotoxicity of SAM were investigated in vitro. Liver function, tumor markers, the extent of solid tumor remission, adverse events, and survival rates were compared between the two groups during a 2-year follow-up period. The diameter of SAM was approximately 180–250 μm and the swelling rate of the microspheres was good, especially at pH 7.4. SAMs did not reduce 3T3-L1 and ECV-304 cell activities and was biocompatible. SAM-TACE was effective in protecting liver function in patients and, when combined with sorafenib, reduced tumor marker levels and cancer volumes, resulting in improved patient survival rates. SAM-TACE combined with sorafenib did not increase the rate of adverse reactions. Therefore, SAM-TACE combined with sorafenib has great potential in the treatment of advanced LC.

Salvianolic acid A exerts a protective effect against sodium laurate induced thromboangiitis obliterans in rats

Purpose: To study the potential therapeutic effects of salvianolic acid A (Sal A) on thromboangiitis obliterans (TAO).
 Methods: An in vitro model of TAO-mimicking endothelial cell damage was established by incubating ECV304 cells with H2O2. An in vivo model of TAO rats was developed via injection of sodium laurate. After treatment with varying doses of Sal A, TAO symptoms were monitored over time and compared. The effects of Sal A on oxidative stress, pyroptosis, inflammation, and thrombosis were assessed using biochemical assays, enzyme-linked immunosorbent assay (ELISA), western blot, and histopathology.
 Results: Sal A significantly alleviated the inhibition of cell viability induced by H2O2 (p < 0.05). The H2O2-induced increases in levels of ROS, IL-1β, and IL-18 in ECV304 cells were significantly decreased by Sal A (p < 0.05). Moreover, Sal A alleviated TAO symptoms in rats. The enhanced levels of IL-1β, IL-18, NLRP3, and active Caspase-1 observed in TAO rats were reduced by Sal A in vivo (p < 0.05). Moreover, Sal A significantly reduced the size of thrombus in TAO rats. In addition, upregulated levels of ROS were significantly inhibited by Sal A (p < 0.05).
 Conclusion: Sal A exerts significant therapeutic effects on TAO. This provides mechanistic insights into the clinical effects of RSMA on TAO patients, which might be beneficial in the development of effective drugs against TAO in the future.

Vitamin D increases the efficacy of cisplatin on bladder cancer cell lines.

1,25(OH)2D3(Calcitriol), which is a broad regulatory molecule, plays a role in changing the efficacy of chemotherapeutic drugs. Cisplatin is one of a current standard chemotherapy regimen for bladder cancer. Increasing the effectiveness of the treatment and reducing the side effects to chemotherapeutics are of great importance in bladder cancer. We aimed to investigate the effect of the combination of cisplatin and calcitriol in order to create a possible advantage in treatment of bladder cancer. T24, ECV-304 and HUVEC cell lines were treated with calcitriol and cisplatin individually and in combination. Dose determination and combination treatments of calcitriol and cisplatin were evaluated using the MTT assay for cytotoxicity analysis on the cells. Annexin V-PI staining method was used for apoptosis determination by flow cytometry. Also the P-gp expression levels were determined by flow cytometry. The combination treatment increased the anti-proliferative efficacy compared to the efficacy in cisplatin alone in T24 cells and reduced the cytotoxicity in the HUVEC healthy cells compared to cisplatin alone. Combination treatment achieved significantly higher apoptosis rate in T24 cells compared with the rates in treatment of cisplatin alone. However apoptosis decreased in HUVEC cell line. P-gp ratios were increased in HUVEC and decreased in T24 cells with combination treatment compared to the numbers in the control cells. The rate of apoptosis and P-gp levels showed no significant change in ECV-304 cells. Our study revealed that the combination of calcitriol and cisplatin allows the use of cisplatin at lower doses in T24 bladder cancer cell line.

Effects of dual-gene modification on biological characteristics of vascular endothelial cells and their significance as reserving cells for chronic wound repair

bFGF is a commonly used and reliable factor for improving chronic wound healing, and hSulf-1 expression is abundant in surrounding cells of chronic wound tissue and vascular endothelial cells, which can reverse the effect of bFGF and inhibit the signalling activity of cell proliferation. In this study, an adenovirus, Ad5F35ET1-bFGF-shSulf1, was designed for establishing the dual-gene modified vascular endothelial cells, which were used as the repair cells for skin chronic wound. Ad5F35ET1-bFGF-shSulf1 infected ECV304 cells in vitro and mediated the overexpression of bFGF and the knockdown of hSulf-1, which effectively activated the AKT and ERK signal transduction pathways, facilitate cell proliferation and migration, with the cell viability to 128.29% at 72 h after infection, compared to 66.65%, 73.74%, 87.63%, 103.14% in the blank control, Ad5F35ET1-EGFP-shNC, Ad5F35ET1-shSulf1, Ad5F35ET1-bFGF groups, respectively. In the rat ear skin injury model, the wound healing was significantly accelerated in the Ad5F35ET1-rbFGF-shrSulf1 group compared to the blank control group (p = 0.0046), Ad5F35ET1-EGFP-shNC group (p = 0.0245), Ad5F35ET1-shrSulf group (p = 0.0426), and Ad5F35ET1-rbFGF group (p = 0.2853). The results demonstrated that this strategy may be a candidate therapy for chronic injury repair.

ENOS Location is a Fundamental Mechanism for Inactivation of Inflammation‐induced Hyperpermeability

Inflammation disrupts the endothelial barrier and increases microvascular permeability (hyperpermeability), leading to tissue edema. Mechanisms for the onset of hyperpermeability have been the focus of many studies. However, the major pathological consequences are related to impairment in terminating hyperpermeability, rather than its onset, and to sustained inflammation. Therefore, we are innovatively studying mechanisms involved in the inactivation of hyperpermeability and restoration of endothelial barrier and normal microvascular permeability. We demonstrated that agonist‐induced translocation of eNOS (endothelial nitric oxide synthase) from cell membrane to cytosol initiates hyperpermeability. We hypothesize that agonist signaling for hyperpermeability also initiates a delayed increase in [cAMP], causing translocation of eNOS and Epac1 (exchange protein activated by cAMP) from cytosol to the cell membrane, which inactivates hyperpermeability. We tested whether platelet‐activating factor (PAF) induced hyperpermeability first and in delayed manner increased cAMP concentration to selectively stimulate Epac1 leading to translocation of eNOS from cytosol back to the cell membrane. In addition, we studied whether vasodilator‐stimulated phosphoprotein (VASP) is involved in the inactivation of hyperpermeability since PAF‐induced hyperpermeability in VASP‐KO endothelial cells remains elevated after Epac1 stimulation. We used human microvascular endothelial cells (HMVEC) and ECV‐304 cells transfected with green fluorescent protein (GFP)‐conjugated eNOS or the constructs GFPeNOSG2A (which anchors eNOS to the cytoplasm, ECV‐GFPeNOS‐G2A) and GFPeNOS‐CAAX (which anchors eNOS to the plasma membrane, ECV‐GFPeNOS‐CAAX) to study the restoration of endothelial barrier after hyperpermeability induced by PAF or vascular endothelial growth factor (VEGF). The time‐correlation between [NO] and [cAMP] in PAF or VEGF‐stimulated HMVEC and ECV‐eNOSGFP indicates that NO increase precedes [cAMP] increase. The increase in [cAMP] in HMVEC and ECV‐eNOSGFP correlates with a close proximity between eNOS, Epac1 and VASP, and both eNOS and Epac1 translocate to the cell membrane. None of these observations occur when eNOS is anchored to the cell membrane or cytosol. Furthermore, stimulation of Epac1 in HMVEC and ECV‐eNOSGFP inactivates their hyperpermeability response to PAF or VEGF. Stimulation of Epac1 in ECV‐GFPeNOS‐CAAX, after PAF application, inhibits hyperpermeability, while Epac1 stimulation does not reduce PAF‐elicited hyperpermeability in ECV‐GFPeNOS‐G2A. Stimulation of Epac1, after application of PAF, in the in vivo male and female hamster cheek pouch prevents hyperpermeability. In conclusion, our results support the concept that eNOS, cAMP, Epac1 and VASP work in synchrony to terminate hyperpermeability by translocating eNOS back to the cell membrane, and the overarching concept that location of eNOS is an important determinant in the regulation of microvascular permeability.

Yoghurt production using pinecone and investigation of the effect of the produced yogurt on ECV304 cell line

OBJECTIVE: The goal of this study was to ferment yoghurt from milk using lactic acid bacteria found in pinecones and to examine the cell viability and lipid peroxidation levels of the fermented yoghurt on Vascular Endothelial Cell Line ECV304.
 METHODS: ECV304 was cultured in vitro. To determine cell viability, various concentration of yogurt extract fermented with pinecone were given to cultured cells using the MTT Assay. At the same doses as the MTT Assay, the Lipid Peroxidation (LPO) Assay was employed to evaluate the malondialdehyde (MDA) levels of cells. Acridine orange/Ethidium Bromide staining technique was applied to detect apoptosis. Gas Chromatography-Mass Spectrometry was used to identify the volatiles in yoghurt fermented with pinecone. RESULTS: All dosages of pinecone fermented yogurt enhanced cell viability of the human healthy vascular endothelial cell line ECV304 and decreased MDA levels, as validated by fluorescence microscopy pictures. The primary essential oils identified in yogurt fermented with pinecone were hexadecanoic acid, eicosanoic acid, stearic acid, and 2-palmitoylglycerol. CONCLUSIONS: In this work, the effects of yogurt fermented with pinecones on human healthy cell lines were examined for the first time. The study discovered that this yoghurt promotes the proliferation of healthy human cells while decreasing oxidative stress in these cells.

Dose-dependent cytotoxic and proliferative effects of Microcystis aeruginosa extract and its fractions on human endothelial cells

Microcystis aeruginosa, which spreads in five continents in the world and reported in drinking water resources in 257 countries, is a dangerous microalgae for human and animal health due to its toxins. The aim of current study was to evaluate the effects of M. aeruginosa extract and its chromatographically separated fractions on human endothelial cells. In this context, crude extract was prepared from M. aeruginosa cultured in BG-11 medium, and it was fractionated by an optimized HPLC method. Algae extract and its six fractions were then analyzed for their cytotoxic effects on ECV304 using MTT assay. The results revealed that algae extract inhibited ECV304 cells by around 72%, a higher percentage than all fractions. The most toxic fraction was the first fraction, which inhibited the cells by 55%. Other fractions, except the third one, were also toxic with 35-40% inhibition percentages. Third fraction and certain doses of some fractions showed proliferative activity on ECV304 cells. These results showed that the activities of the total extract and its fractions in promoting or inhibiting cell proliferation varied depending on not only the content but also the treatment dose.

The role of pumpkin pulp extract carotenoids against mycotoxin damage in the blood brain barrier in vitro.

Some mycotoxins such as beauvericin (BEA), ochratoxin A (OTA), and zearalenone (ZEA) can cross the blood brain barrier, which is why we tested the anti-inflammatory action of a pumpkin carotenoid extract (from the pulp) against these mycotoxins and their combinations (OTA+ZEA and OTA+ZEA+BEA) on a blood brain barrier model with co-cultured ECV304 and C6 cells using an untargeted metabolomic approach. The cells were added with mycotoxins at a concentration of 100 nmol/L per mycotoxin and pumpkin carotenoid extract at 500 nmol/L. For control we used only vehicle solvent (cell control) or vehicle solvent with pumpkin extract (extract control). After two hours of exposure, samples were analysed with HPLC-ESI-QTOF-MS. Metabolites were identified against the Metlin database. The proinflammatory arachidonic acid metabolite eoxin (14,15-LTE4) showed lower abundance in ZEA and BEA+OTA+ZEA-treated cultures that also received the pumpkin extract than in cultures that were not treated with the extract. Another marker of inflammation, prostaglandin D2-glycerol ester, was only found in cultures treated with OTA+ZEA and BEA+OTA+ZEA but not in the ones that were also treated with the pumpkin extract. Furthermore, the concentration of the pumpkin extract metabolite dihydromorelloflavone significantly decreased in the presence of mycotoxins. In conclusion, the pumpkin extract showed protective activity against cellular inflammation triggered by mycotoxins thanks to the properties pertinent to flavonoids contained in the pulp.

Shenlian extract attenuates TNF-α-induced ECV304 injury by regulating Nrf2/Keap1 signaling pathway

This study aimed to investigate the effect and the possible mechanism of Shenlian( SL) extract on tumor necrosis factor-α( TNF-α)-induced ECV304 injury. After the establishment of TNF-α-induced ECV304 cells injure model,MTT assay was used to detect cell viability and the level of reactive oxygen species( ROS) was measured by flow cytometry. The contents of superoxide dismutase( SOD),malondialdehyde( MDA),nitric oxide( NO),endothelin-1( ET-1) and interleukin-1β( IL-1β) in the supernatant were detected by biochemical method and enzyme linked immunosorbent assay( ELISA). The expression levels of apoptosis-related proteins B-lymphoma-2 gene( Bcl-2),Bcl-2 associated X protein( Bax),caspase-3,caspase-9 and nuclear factor E2 associated factor2( Nrf2)/Kelch like epichlorohydrin associated protein-1( Keap1) signaling pathway related proteins Nrf2,Keap1,quinone oxidoreductase( NQO1) and heme oxygenase 1( HO-1) were detected by Western blot. The results showed that 50 μg·L-1 TNF-α significantly damaged ECV304 cells,induced the impairment of cell viability( P<0. 01),the increase of ROS production,the decrease of SOD activity,and the increase of MDA,NO,ET-1 and IL-1β( P<0. 01),meanwhile,it caused the up-regulation of Keap1,caspase-9 and Bax protein expression,and down-regulation of NQO1 and Bcl-2 protein expression( P<0. 05) compared with the control group.Compared with the model group,SL extract reduced the damage of ECV304 cells induced by TNF-α,improved cell viability,reduced ROS production,increased SOD activity and decreased MDA,NO,ET-1,IL-1β content( P<0. 01 or P<0. 05). In addition,SL extract also down-regulated the protein expression levels of Keap1,caspase-3,caspase-9 and Bax,and increased the protein expressions of Nrf2,NQO1,HO-1 and Bcl-2( P<0. 01 or P<0. 05). The above results indicate that SL extract can provide protective effect on ECV304 cells injury induced by TNF-α,alleviate oxidative stress injury,inflammation and apoptosis,and its mechanism may be related to regulating Nrf2/Keap1 signaling pathway.

Plasmatic and cell-based enhancement by microparticles originated from platelets and endothelial cells under simulated in vitro conditions of a dilutional coagulopathy

BackgroundAggressive fluid management and other external factors may lead to hypothermia, acidosis and hemodilution (defined as Lethal Triad, LT) contributing to a trauma-induced coagulopathy (TIC) that worsens patients’ outcomes. Procoagulant microparticles (MP) are crucial players at the interface of cellular and plasmatic coagulation. However, their functions remain largely unexplored. This study aimed to characterize effects of MP subtypes and concentrations on functional coagulation under in vitro simulated conditions.MethodsBlood from eleven volunteers were collected to simulate in vitro conditions of hemodilution (HD) and LT, respectively. HD was induced by replacing a blood volume of 33% by crystalloids and for LT, samples were further processed by reducing the temperature to 32 °C and lowering the pH to 6.8. MP were obtained either from platelet concentrates (platelet-derived MP, PDMP) or from cell culture (ECV304 cells for endothelial-derived MP, EDMP) by targeted stimulation. After introducing MP to in vitro conditions, we measured their concentration-dependent effects (1.000, 10.000 and 15.000 MP/μl blood) on coagulation compared to whole blood (WB). For each condition, coagulation was characterized by flow cytometric platelet activation and by quantification of fibrin clot propagation using Thrombodynamics® technology.ResultsMP originated from platelets and endothelial cells affected blood coagulation in a concentration-dependent manner. Particularly, high PDMP quantities (10.000 and 15.000 PDMP/μl blood) significantly induced platelet activation and fibrin clot growth and size in HD conditions. In LT conditions as well, only high PDMP concentration induced platelet activation, clot growth and size. In contrast, EDMP did not induce platelet activation, but resulted in enhanced formation of spontaneous clots, irrespective of simulated condition. With increasing EDMP concentration, the time until the onset of spontaneous clotting decreased in both HD and LT conditions.DiscussionThe study demonstrates an essential role of MP within the coagulation process under simulated coagulopathic conditions. PDMP affected platelets promoting clot formation likely by providing a surface enlargement. EDMP presumably affected clotting factors of the plasmatic coagulation resulting in an increased formation of spontaneous clots.ConclusionUnder simulated conditions of a dilutional coagulopathy, MP from different cellular origin indicate a divergent but both procoagulant mechanism within the coagulation process.

Comparative study of dehydrated papaya (Vasconcellea pubescens) by different drying methods: quality attributes and effects on cells viability

The effect of different drying methods on sugar content and amino acid content, color and non-enzymatic browning of Chilean papaya (Vasconcellea pubescens) slices was investigated. The obtained fruit extracts were tested in cell viability assays on human endothelial ECV-304 cells. Drying techniques included freeze-, vacuum-, solar-, convective- and infrared-drying. Consistently, infrared-dried papaya had a lower sugar content and a higher non-enzymatic browning intensity than papaya dehydrated by the other methods. All dried samples were lighter in color with a lower yellow intensity compared to fresh papaya. The amino acid lysine was the most abundant in the infrared-dried sample. On the other side, the methods that employed vacuum, increased their cellular viability. Based on these results, operational parameters during drying processes should be considered to preserve, on one hand, product quality attributes, while, on the other hand, increasing cell viability.

MiR-100 alleviates the inflammatory damage and apoptosis of H2O2-induced human umbilical vein endothelial cells via inactivation of Notch signaling by targeting MMP9.

Thromboangiitis obliterans is a nonatherosclerotic segmental inflammatory disease, and miR-100 plays an anti-inflammatory role in chronic inflammation. Therefore, we hypothesized that miR-100 might alleviate the inflammatory damage and apoptosis of H2O2-induced ECV304 cells and aimed to investigate the relationship between miR-100 and thromboangiitis obliterans and the related molecular mechanism. Cell counting kit-8 was used to detect cell viability, and the expression of inflammatory factors and oxidative stress was measured by ELISA. TUNEL assay was used to detect the apoptosis of human umbilical vein endothelial cells after induction by H2O2. Furthermore, the interaction between miR-100 and matrix metalloproteinase-9 was verified by dual-luciferase assay. Quantitative reverse transcription polymerase chain reaction and western blot were used to detect the expression of the adhesion factors, apoptosis-related proteins and Notch pathway-related protein. The results revealed that miR-100 was decreased in H2O2-induced human umbilical vein endothelial cells. Overexpression of miR-100 attenuated inflammatory response and cell apoptosis in H2O2-induced human umbilical vein endothelial cells. The overexpression of miR-100 inhibited matrix metalloproteinase-9 expression in H2O2-induced human umbilical vein endothelial cells. miR-100 inhibited H2O2-induced human umbilical vein endothelial cell inflammation, oxidative stress, and cell apoptosis via inactivation of Notch signaling by targeting matrix metalloproteinase. Our study demonstrated that miR-100 reduced the inflammatory damage and apoptosis of H2O2-induced human umbilical vein endothelial cells via inactivation of Notch signaling by targeting matrix metalloproteinase. These findings suggested that miR-100 might be a novel therapeutic target for the prevention of thromboangiitis obliterans.

Evaluation of Fluorescent Cu2+ Probes: Instant Sensing, Cell Permeable Recognition and Quantitative Detection.

By incorporating a rhodamine spirolactam structure as the recognition site for Cu2+, two novel probes were synthesized through a connection of rhodamine 6G acylhydrazine and 5-formyl-6-hydroxyl-4-methylcoumarin/2,4-dihydroxybenzaldehyde. In the recognition process of probes towards Cu2+, the spirolactam ring exhibited opening and closing, accompanying an instant and specific change in fluorescence and in color, which could also achieve a naked-eye and semiquantitative recognition of aqueous Cu2+ besides the fluorescent Cu2+ detection method. Fluorescent analyses and ECV304 cell imaging further revealed the probes’ good optical stability, instant response, low toxicity, and membrane permeability, which offers future possibilities for the probes’ instant detection and the real-time tracking of Cu2+ in biological systems.

Classification of the mechanism of toxicity as applied to human cell line ECV304

The objective of this study was to identify the pattern of cytotoxicity testing of the human cell line ECV304 using three techniques of an ensemble learning algorithm (bagging, boosting and stacking). The study of cell morphology of ECV304 cell line was conducted using impedimetric measurement. Three types of toxins were applied to the ECV304 cell line namely 1 mM hydrogen peroxide (H2O2), 5% dimethyl sulfoxide and 10 μg Saponin. The measurement was conducted using electrodes and lock-in amplifier to detect impedance changes during cytotoxicity testing within a frequency range 200 and 830 kHz. The results were analysed, processed and extracted using detrended fluctuation analysis to obtain characteristics and features of the cells when exposed to the each of the toxins. Three ensemble algorithms applied showed slightly different results on the performance for classifying the data set from the feature extraction that was performed. However, the results show that the cell reaction to the toxins could be classified.

Suppression of Urokinase-Type Plasminogen Activator Receptor by Docosahexaenoic Acid Mediated by Heme Oxygenase-1 in 12-O-Tetradecanoylphorbol-13-Acetate-Induced Human Endothelial Cells.

Urokinase-type plasminogen activator receptor (uPAR) plays a crucial role in inflammation and tumor metastasis. Docosahexaenoic acid (DHA), a representative omega-3 polyunsaturated fatty acid, has been shown to exhibit anti-inflammatory and anti-tumor properties. However, the mechanism by which DHA negatively regulates uPAR expression is not yet understood. The aim of this study was to investigate the effect of DHA on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced uPAR expression and potential role of heme oxygenase-1 (HO-1) in DHA-induced inhibition of uPAR in human endothelial ECV304 cells. Results showed that TPA induced uPAR expression in a time dependent manner, while DHA inhibited uPAR expression in a concentration-dependent manner. Moreover, treatment with DHA induced HO-1 expression in a time- and concentration-dependent manner. In addition, DHA-induced inhibition of uPAR expression and cell invasion in TPA-stimulated cells was reversed by si-HO-1 RNA. Induction of HO-1 by ferric protoporphyrin IX (FePP) inhibited TPA-induced uPAR expression, and this effect was abolished by treatment with the HO-1 inhibitor tin protoporphyrin IX (SnPP). Additionally, carbon monoxide, an HO-1 product, attenuated TPA-induced uPAR expression and cell invasion. Collectively, these data suggest a novel role of DHA-induced HO-1 in reducing uPAR expression and cell invasion in human endothelial ECV304 cells.

Aqueous Zn2+ analysis: Specific recognition and instant imaging by Schiff base fluorescent probes

Two Schiff base fluorescence probes for aqueous Zn2+ were synthesized by introducing salicylaldehyde benzoyl hydrazine as the recognition group and C=N as the characteristic fluorescence chromophore. Spectroscopic analyses, mainly FS, illuminated the probes’ highly specific recognition towards Zn2+ over other metal ions, accompanied by steady optical properties at a wide pH range of 6.0 to 9.0. Moreover, cytotoxicity and fluorescence imaging in living ECV304 cells identified the probes’ instant response and membrane permeability. These results predicted the probes’ future applications for instant Zn2+ detection in a clinical diagnosis and dynamic analysis of Zn2+ in biological systems.