Journal: Stem Cells International

Article Title: Disrupted Endothelial Cell Layer and Exposed Extracellular Matrix Proteins Promote Capture of Late Outgrowth Endothelial Progenitor Cells

doi: 10.1155/2016/1406304

Figure Lengend Snippet: Representative phase-contrast live cell images of interaction of LO-EPC with 3 × 10 4 subconfluent (a) and 6 × 10 4 confluent (b) HAEC monolayers under shear stress 0.7 dyn/cm 2 . 4 × 10 5 LO-EPC were perfused for 4 min. Arrows indicated adherent LO-EPC which only adhered paracellularly at junctional regions of discontinuity between two cells in subconfluent HAEC monolayer. Scale bar 30 μ m. After adhesion LO-EPC spread and establish cell-cell interaction (c). LO-EPC were labelled with DiI-Ac-LDL shown red. Scale bar 60 μ m. VE-Cadherin staining revealed the formation of lateral junction between LO-EPC and HAEC (d). LO-EPC were labelled with DiI-Ac-LDL shown red and VE-Cadherin expression shown green.

Article Snippet: To label LO-EPC with Dil-Ac-LDL, LO-EPC were incubated with 10 μ g/mL of Dil-Ac-LDL (Molecular Probes, Invitrogen) for 1 hour at 37°C and then washed twice with PBS.

Techniques: Staining, Expressing

Journal: Stem Cells International

Article Title: Disrupted Endothelial Cell Layer and Exposed Extracellular Matrix Proteins Promote Capture of Late Outgrowth Endothelial Progenitor Cells

doi: 10.1155/2016/1406304

Figure Lengend Snippet: LO-EPC (4 × 10 5 ) were perfused over fibronectin-coated Ibidi slides and time lapse imaging was used to visualise and record LO-EPC adhesion. Representative still images show three adherent cells after initial capture (a, b) after 4 min of perfusion (0.7 dyn/cm 2 ). Scale bar represents 30 μ m. Adherent LO-EPC form lateral adhesion junction (c). LO-EPC labelled with DiI-Ac-LDL shown red and VE-Cadherin expression shown green. Scale bar 20 μ m. 24 hours after adhesion to fibronectin, LO-EPC spread and proliferate (d). Scale bar represents 60 μ m. The adherent LO-EPC effectively cover a fibronectin-coated surface (e). Scale bar represents 110 μ m.

Article Snippet: To label LO-EPC with Dil-Ac-LDL, LO-EPC were incubated with 10 μ g/mL of Dil-Ac-LDL (Molecular Probes, Invitrogen) for 1 hour at 37°C and then washed twice with PBS.

Techniques: Imaging, Expressing

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Effects of myrAkt on nLDL- and oxLDL-induced apoptosis in wild type EPC. a Effects of Ad-DLE1 (control transduction) and Admiralty (constitutively active Akt transduction) on p-Akt, total Akt, p-GSK3α, total GSK3α and β-actin expression in wild-type EPC. b EPC cultures, transducer with either Ad-DLE1 (control; empty vector) or Ad-myrAkt (constitutively active Akt) were exposed to nLDL (10 μg/ml) or oxLDL (10 μg/ml), or oxLDL (10 μg/ml) in serum-free medium. Apoptosis was determined by TUNEL staining. Data are expressed as the percentage of TUNEL positive cells with respect to the total number of cells (DAPI staining) per high power field. Data are presented as mean ± SD, n = 3; + p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: Transduction, Expressing, Plasmid Preparation, TUNEL Assay, Staining

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Effects of SOD, L -NAME, epicatechin and FeTTPs on the ratio of p-Akt/total Akt in wild-type EPC. EPC cultures were exposed to nLDL (10 μg/ml) or oxLDL (10 μg/ml), or oxLDL (10 μg/ml) in the presence of SOD (100 U/ml), L -NAME (0.5 m M ), epicatechin (100 μ M ) or FeTPPs (2.5 μ M ) in serum-free medium. A control well was treated with serum-free medium alone. Density of the Akt and p-Akt bands was first corrected with respect to β-actin (not shown) and then the ratio of p-Akt/total Akt expression calculated. The ratio in the control group was assigned a value of 1.0. Data are presented as mean ± SD, n = 3; + p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: Expressing

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Effects of nLDL and oxLDL on apoptosis in wild-type EPC. EPC cultures were exposed to nLDL (10 μg/ml) or oxLDL (1–50 μg/ml) in serum-free medium. The control well was treated with medium alone. a Percentage of apoptotic cells determined by TUNEL staining. Data are presented as mean ± SD, n = 3 animals; * p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: TUNEL Assay, Staining

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Effects of nLDL and oxLDL on reactive oxygen species in wild-type EPC. EPC cultures were exposed to nLDL (10 μg/ml), oxLDL (10 μg/ml), oxLDL (10 μg/ml) + SOD (100 U/ml) or oxLDL (10 μg/ml) + L -NAME (0.5 m M ) in serum-free medium. The control well was treated with medium alone. a Quantification of DCF and DHE staining in EPC. Data are presented as mean ± SD, n = 3; + p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: Staining

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Effects of SOD, L -NAME, epicatechin and FeTTPs on oxLDL-induced apoptosis in wild-type EPC. EPC cultures were exposed to nLDL (10 μg/ml) or oxLDL (10 μg/ml), or oxLDL (10 μg/ml) in the presence of SOD (100 U/ml), L -NAME (0.5 m M ), epicatechin (100 μ M ) or FeTPPs (2.5 μ M ) in serum-free medium. A control well was treated with serum-free medium alone. Apoptosis was determined by TUNEL staining. Data are expressed as the percentage of TUNEL positive cells with respect to the total number of cells (DAPI staining) per high power field. Data are presented as mean ± SD, n = 3; + p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: TUNEL Assay, Staining

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Effects of SOD, L -NAME, epicatechin and FeTPPs on nitrotyrosine expression and the ratio of the PI3K p85 and p110 subunits in wild-type EPC. EPC cultures were exposed to nLDL (10 μg/ml) or oxLDL (10 μg/ml), or oxLDL (10 μg/ml) in the presence of SOD (100 U/ml), L -NAME (0.5 m M ), epicatechin (100 μ M ) or FeTPPs (2.5 μ M ) in serum-free medium. A control well was treated with serum-free medium alone. a The p110 Akt subunit was isolated by immunoprecipitation; the expression of p110 and nitrotyrosine was determined. Note the essential absence of nitrotyrosine. This set of Western blots is representative of 3 separate experiments. b The p85 Akt subunit was isolated by immunoprecipitation; the expression of p110 and nitrotyrosine were determined. Density of the nitrotyrosine and PI3K p85 subunit bands were first corrected with respect to β-actin and then the ratio of nitrotyrosine/p85 expression calculated. The ratio of the control group was assigned a value of 1.0. Data are presented as mean ± SD, n = 3; + p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: Expressing, Isolation, Immunoprecipitation, Western Blot

Journal: Journal of Vascular Research

Article Title: Oxidized Low-Density Lipoprotein Induces Apoptosis in Endothelial Progenitor Cells by Inactivating the Phosphoinositide 3-Kinase/Akt Pathway

doi: 10.1159/000313879

Figure Lengend Snippet: Time- and dose-response relationships between oxLDL and the expression of Akt and p-Akt in wild-type EPC. a EPC cultures were exposed to nLDL (10 μg/ml) or oxLDL (1–50 μg/ml) in serum-free medium; the control well was treated with serum-free medium alone. Density of the Akt and p-Akt bands were first corrected with respect to β-actin (not shown) and then the ratio of p-Akt/total Akt expression calculated. The ratio of the wild-type group was assigned a value of 1.0. Data are presented as mean ± SD, n = 3; ** p

Article Snippet: After 7 days the identity of adherent cells as EPC was confirmed in some wells by incubation with 2.4 μg/ml Dil-Ac-LDL for 1 h, followed by fixation in 2% paraformaldehyde and counterstaining with FITC-labeled lectin.

Techniques: Expressing