Journal: Nature Communications

Article Title: Soluble E-cadherin promotes tumor angiogenesis and localizes to exosome surface

doi: 10.1038/s41467-018-04695-7

Figure Lengend Snippet: sE-cad-positive exosomes activate the NFκB pathway. a Top network identified by IPA. Gene signatures of HUVEC incubated with IgG pre-treated exosomes and HECD-1 pre-treated exosomes. b NFκB reporter assay of HUVEC treated with exosomes (25 μg mL −1 ) in the presence or absence of E-cadherin neutralizing antibodies, HECD-1 (100 μg mL −1 ) or VE-cadherin neutralizing antibodies, BV9 (10 μg mL −1 ). c Western blot analysis of the subcellular localization of NFκB subunits in HUVEC treated with exosomes (25 μg mL −1 ) in the presence or absence of E-cadherin neutralizing antibodies, HECD-1 (100 μg mL −1 ). d Tube formation assay of HUVEC treated with exosomes (25 μg mL −1 ) in the presence or absence of Bay11-7082 (Bay) (10 μM). Upper: Representative images of HUVEC tube formation assay. Lower: Quantification of the percentage change of the number of branching points. Bar, 100 μm. e Coimmunoprecipitation of NFκB subunits and β-catenin (β-cat) in HUVEC with or without exosomes treatment (25 μg mL −1 ). f NFκB reporter assay of HUVEC transiently transfected with non-specific (NS) siRNA or β-catenin (β-cat) siRNA (20 nM) for 24 h, treated with exosomes (25 μg mL −1 ). TCF reporter assay of HUVEC transiently transfected with NS siRNA or p105/p50 and p65 siRNA (20 nM) for 24 h, treated with exosomes (25 μg mL −1 ). g Western blot analysis of the β-catenin (β-cat) and NFκB subunits in HUVEC treated with exosomes (25 μg mL −1 ) for the indicated period of time. Upper: Representative images of nuclear fractions. Lower: Densitometry of β-catenin (β-cat) and p65. h Tube formation assay of HUVEC transiently transfected with non-specific (NS) siRNA, β-catenin siRNA, or p105/p50 siRNA and p65 siRNA (20 nM) for 24 h with or without exosomes treatment (25 μg mL −1 ). Upper: Representative images of HUVEC tube formation assay. Lower: Quantification of the percentage change of the number of branching points. Bar, 100 μm. n = 3 per group, all experiments were repeated three times. Error bar indicates SD of the mean. * P < 0.05 versus untreated control using one-way analysis of variance followed by Tukey’s least significant difference post hoc test

Article Snippet: Equal amounts of protein (1 mg) were precleared with protein A/G PLUS agarose beads (Santa Cruz Biotechnology) for 2 h. Supernatant was then incubated with anti-p65 antibodies or mouse IgG at 4 °C overnight.

Techniques: Incubation, Reporter Assay, Western Blot, Tube Formation Assay, HUVEC Tube Formation Assay, Transfection, Control

Journal: PLoS ONE

Article Title: Expansion of Endothelial Progenitor Cells in High Density Dot Culture of Rat Bone Marrow Cells

doi: 10.1371/journal.pone.0107127

Figure Lengend Snippet: A, Flow cytometric analysis showed that the majority of cells in regular density (RD) culture became larger after their expansion, whereas the cells in high density (HD) culture maintained the small size of their parental cells (P0). B, Cells from high density culture were analyzed further by gating based on different cell sizes. The small cells showed higher expression levels of EPC related markers than those of the large cells. C, In high density culture, the majority of cells positive for KDR, CD34 and DiI were also positive for CD45. D, In vitro angiogenesis assay of CD45 + and CD45 − cells sorted from high density culture. E, CD45 + , but not CD45 − , cells from high density culture differentiated into endothelial cells that were able to uptake DiI-ac-LDL and express vWF and eNOs. Scale bars, 100 µm. F, Percentages of endothelial cell marker-positive cells in CD45 + and CD45 − populations after 14 days of induction (n = 3). *p<0.05.

Article Snippet: To detect the angiogenic inductive potential of the supernatants, 2×10 4 human umbilical vein endothelial cells (HUVECs; Sciencell, Carlsbad, CA, USA) were seeded onto a Matrigel-coated 24-well plate in 500 μl of supernatants from regular or high density cultures.

Techniques: Expressing, In Vitro, Angiogenesis Assay, Marker

Journal: PLoS ONE

Article Title: Expansion of Endothelial Progenitor Cells in High Density Dot Culture of Rat Bone Marrow Cells

doi: 10.1371/journal.pone.0107127

Figure Lengend Snippet: A and B, Confocal images revealed that some of high density cultured cells (CM-DiI + , white arrows) participated in neovascularization by direct differentiation into endothelial cells (ILB4 + ). C, CM-DiI + cells (white arrows) close to capillaries, but negative for ILB4 staining, might support vessel formation via paracrine effect of released pro-angiogenic growth factors. D, The percentages of endothelial and non-endothelial differentiation of injected cells, and the percentage of vessels derived from donor and recipient (n = 3). *p<0.05.

Article Snippet: To detect the angiogenic inductive potential of the supernatants, 2×10 4 human umbilical vein endothelial cells (HUVECs; Sciencell, Carlsbad, CA, USA) were seeded onto a Matrigel-coated 24-well plate in 500 μl of supernatants from regular or high density cultures.

Techniques: Cell Culture, Staining, Injection, Derivative Assay