human coronary artery endothelial cells hcaecs  (Cell Applications Inc)

Journal: PLoS ONE

Article Title: Endothelial cell activation is attenuated by everolimus via transcriptional and post-transcriptional regulatory mechanisms after drug-eluting coronary stenting

doi: 10.1371/journal.pone.0197890

Figure Lengend Snippet: Immunohistochemical staining and analysis of NF-κB activation in TNF-α and everolimus treated endothelial cells. HCAEC cells were treated for 1 hour with MesoEndo Medium (Baseline), 100 ng/ml TNF-α with or without 0.5 μM everolimus or cytokine with the solvent of everolimus (TNF-α+DMSO). Nuclear localization of the NF-κB p65 subunit was monitored by immunostaining. Green: p65 staining; blue: cell nuclei. Scale bar: 20 μm (A). Ratio of the fluorescence intensity of the NF-κB immunostaining in cell nuclei and cytosol was analyzed (B). Mean ± SEM, n = 6-8/group.

Article Snippet: Culturing endothelial cells with or without everolimus Human coronary artery endothelial cells (HCAEC, Cell Applications Inc, San Diego, CA, USA) were cultured in ready-to-use MesoEndo Cell Growth Medium (Cell Applications) at 37°C, 5% CO2 .

Techniques: Immunohistochemistry, Staining, Activation Assay, Immunostaining, Fluorescence

Journal: PLoS ONE

Article Title: Endothelial cell activation is attenuated by everolimus via transcriptional and post-transcriptional regulatory mechanisms after drug-eluting coronary stenting

doi: 10.1371/journal.pone.0197890

Figure Lengend Snippet: Quantification of TNF-α induced miR-155 and miR-181b levels with the analysis of their precursors in the presence of everolimus upon EC inflammation in vitro . HCAECs were treated by recombinant TNF-α for 1–4 hours to analyze miR-181b expression along with the inflammation-specific miR-155. First, miR-155 was elevated by TNF-α compared to untreated sample, however, everolimus caused significantly decreased miR-155 levels (A), while miR-181b was downregulated by the inflammatory stimulus and the treatment with everolimus restored their expression in both EC cultures (B). Levels of pre- and pri-miRNA were altered in the same manner as seen in mature miR-155 (C, E) and miR-181b (D, F), respectively. Mean ± SEM, n = 4-8/group.

Article Snippet: Culturing endothelial cells with or without everolimus Human coronary artery endothelial cells (HCAEC, Cell Applications Inc, San Diego, CA, USA) were cultured in ready-to-use MesoEndo Cell Growth Medium (Cell Applications) at 37°C, 5% CO2 .

Techniques: In Vitro, Recombinant, Expressing

Journal: PLoS ONE

Article Title: Endothelial cell activation is attenuated by everolimus via transcriptional and post-transcriptional regulatory mechanisms after drug-eluting coronary stenting

doi: 10.1371/journal.pone.0197890

Figure Lengend Snippet: Analysis of E-selectin and VCAM-1 expression at mRNA and protein levels in HCAECs after TNF-α stimulation. HCAEC cells were treated with recombinant TNF-α (100 ng/mL) for 1–24 hours to generate cellular inflammatory conditions. Elevated E-selectin (SELE) (A) and VCAM-1 mRNA levels (B) induced by TNF-α already after 1 hour were downregulated by everolimus in HCAECs in vitro . In parallel, soluble E-selectin (C) and VCAM-1 concentrations (D) were measured by ELISA in the supernatants of ECs and were also significantly decreased after 4 and 24 hours, respectively. Mean ± SEM, n = 4-8/group.

Article Snippet: Culturing endothelial cells with or without everolimus Human coronary artery endothelial cells (HCAEC, Cell Applications Inc, San Diego, CA, USA) were cultured in ready-to-use MesoEndo Cell Growth Medium (Cell Applications) at 37°C, 5% CO2 .

Techniques: Expressing, Recombinant, In Vitro, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: Endothelial cell activation is attenuated by everolimus via transcriptional and post-transcriptional regulatory mechanisms after drug-eluting coronary stenting

doi: 10.1371/journal.pone.0197890

Figure Lengend Snippet: Everolimus decreased EC inflammation via lowered IL-1β and IL-6 expression in ECs. HCAECs were treated with TNF-α (100 ng/mL) with or without everolimus (0.5 μM) for 1 and 4 hours, and then IL-1β and IL-6 mRNA levels were quantified by RT-qPCR. This effect was observed already after 1 hour (A, C), while was more pronounced after 4 hours (B, D). Mean ± SEM, n = 4-8/group.

Article Snippet: Culturing endothelial cells with or without everolimus Human coronary artery endothelial cells (HCAEC, Cell Applications Inc, San Diego, CA, USA) were cultured in ready-to-use MesoEndo Cell Growth Medium (Cell Applications) at 37°C, 5% CO2 .

Techniques: Expressing, Quantitative RT-PCR

Journal: PLoS ONE

Article Title: Endothelial cell activation is attenuated by everolimus via transcriptional and post-transcriptional regulatory mechanisms after drug-eluting coronary stenting

doi: 10.1371/journal.pone.0197890

Figure Lengend Snippet: Analysis of transcriptional regulation of SELE and VCAM-1 genes in unstimulated and TNF-α-treated HUVECs using publicly available ChIP-seq data sets and the measurement of eRNA expression at two selected enhancers including SELE_-11Kb and VCAM1_-10Kb in HCAECs. The p65, RNAPII, H3K27Ac and H3K4m3-specific ChIP-seq signals at the selected VCAM1 and SELE-associated enhancers were visualized by the Integrative Genomics Viewer (A). Accordingly, HCAECs were then treated with TNF-α (100 ng/mL) with or without everolimus (0.5 μM) for 1 hour, and one-one eRNA expression at SELE_-11Kb (B) and VCAM1_-10Kb eRNAs (C) were quantified by RT-qPCR. TNF-α-augmented expression of eRNAs was significantly reduced by everolimus treatment. Mean ± SEM, n = 4-8/group.

Article Snippet: Culturing endothelial cells with or without everolimus Human coronary artery endothelial cells (HCAEC, Cell Applications Inc, San Diego, CA, USA) were cultured in ready-to-use MesoEndo Cell Growth Medium (Cell Applications) at 37°C, 5% CO2 .

Techniques: Chromatin Immunoprecipitation, Expressing, Quantitative RT-PCR

Journal: PLoS ONE

Article Title: Endothelial cell activation is attenuated by everolimus via transcriptional and post-transcriptional regulatory mechanisms after drug-eluting coronary stenting

doi: 10.1371/journal.pone.0197890

Figure Lengend Snippet: Schematic figure about the model to demonstrate the regulatory mechanisms of everolimus on E-selectin (SELE) and VCAM-1 expression. Everolimus decreases EC activation via suppressing the NF-κB pathway with decreased p65 translocation into cell nuclei causing the modulation of E-selectin and VCAM-1 expression as well as miR-181b level at transcriptional and post-transcriptional level, respectively. EC: endothelial cell, TNF-α: tumor necrosis factor alpha, SELE: E-selectin, VCAM-1: vascular cell adhesion molecule-1, RISC: RNA-induced silencing complex.

Article Snippet: Culturing endothelial cells with or without everolimus Human coronary artery endothelial cells (HCAEC, Cell Applications Inc, San Diego, CA, USA) were cultured in ready-to-use MesoEndo Cell Growth Medium (Cell Applications) at 37°C, 5% CO2 .

Techniques: Expressing, Activation Assay, Translocation Assay

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Moderate‐ and High‐Intensity Exercise Improves Lipoprotein Profile and Cholesterol Efflux Capacity in Healthy Young Men

doi: 10.1161/JAHA.121.023386

Figure Lengend Snippet: Exercise increases ABCA1‐mediated cholesterol efflux and improves the anti‐inflammatory properties of HDLs. ABCA1‐specific cholesterol efflux at baseline and after moderate‐ and high‐intensity exercise. Chinese hamster ovary cells with tetracycline‐inducible expression of human ABCA1 were loaded with 3 H‐cholesterol and incubated for 4 hours in the absence or presence of apoB‐depleted serum as described in the Methods. The percentage of efflux was calculated as radioactivity in the medium relative to the total radioactivity (cells+medium). ABCA1‐specific cholesterol efflux was calculated as the difference in efflux between cells incubated with and without tetracycline and normalized to a standard sample of apoB‐depleted serum (n=51) ( A ). ICAM‐1 and VCAM‐1 expression in TNF‐α activated HCAECs at baseline and after moderate‐ and high‐intensity ( B and C ). HCAECs were preincubated for 16 hours with ultracentrifugally isolated HDLs at baseline and after moderate‐ and high‐intensity exercise, then activated for 5 hours with TNF‐α. Expression of ICAM‐1 (n=19) ( B ) and VCAM‐1 (n=19) ( C ) was quantified by flow cytometry. Data represent the mean±SEM of triplicate measurements. Repeated measures ANOVA was conducted with Bonferroni post hoc pairwise comparison testing for multiple comparisons. *** P

Article Snippet: Human coronary artery endothelial cells (HCAECs) (Cell Applications Inc., San Diego CA) were grown to 90% confluence in 5% CO2 in Meso Endo Growth Medium (Cell Applications) and seeded onto 24‐well tissue culture plates (5×104 cells/well) in Meso Endo Growth Medium (500 µL).

Techniques: Expressing, Incubation, Radioactivity, Isolation, Flow Cytometry

Journal: PLoS ONE

Article Title: Distinct Effects of Unfractionated Heparin versus Bivalirudin on Circulating Angiogenic Peptides

doi: 10.1371/journal.pone.0034344

Figure Lengend Snippet: Levels of angiogenic peptides from human coronary artery endothelial cells treated with unfractionated heparin or bivalirudin. A) Conditioned media levels of sFlt1, PlGF, VEGF and sEng 30 minutes after treatment with increasing concentrations of UFH. (p

Article Snippet: Circulating Angiogenic Peptides in vitro Human coronary artery endothelial cells (HCAEC; Cell Applications Inc., San Diego, CA, USA) were cultured to near confluence using normal humidified tissue culture incubators with 5% CO2.

Techniques: