Journal: Cells
Article Title: ADAMTS13 Improves Endothelial Function and Reduces Inflammation in Diabetic Retinopathy.
doi: 10.3390/cells14020085
Figure Lengend Snippet: Figure 11. Inhibition of endothelial cell migration by ADAMTS13. (A,B) Confluent monolayers of overnight starved HRMECs were scratched with sterile micropipette tips and monolayer regeneration was microscopically monitored, subject to various treatments. In one set of experiments, the cultures were pretreated with a dilution medium or ADAMTS13 (60, 200, or 600 ng/mL) for 1 h, followed by stimulation with vascular endothelial growth factor (VEGF) (10 ng/mL) for 16 h. Two independent experiments were performed in duplicates and two-to-three independent field images were taken for the migration analysis with the Image J software (summarized in (B)). In panel (A), representative images illustrate the effect of ADAMTS13, at a dose of 600 ng/mL, on VEGF-induced cell migration. (C) In a second set of experiments, endothelial cell migration through 8 µm pores of polyethylene terephthalate (PET) membranes in response to VEGF (10 ng/mL) with or without pretreatment with ADAMTS13 (6 to 600 ng/mL) was analyzed with the xCELLigence instrument (three or four independent experiments in duplicates). Results are expressed as means ± standard deviation. One- way ANOVA and independent t-tests were used for comparisons among five groups and between two groups, respectively. * p < 0.05 compared with values obtained from untreated cells. # p < 0.05 compared with values obtained from cells treated with VEGF only.
Article Snippet: The immunodetection of specific molecules was conducted with the following reagents and conditions: VWF with a mouse monoclonal anti-VWF antibody (1:1000, sc-365712, Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA), CD41 with a mouse monoclonal antiCD41 antibody (1:1000, sc-365938, Santa Cruz Biotechnology Inc.), β-catenin with a goat polyclonal anti-ß-catenin antibody (1:1000, AF1329, R&D system, Minneapolis, MN, USA), ADAMTS13 with a rabbit monoclonal anti-ADAMTS13 antibody (1:1000, NBP3-16038, Novus Biologicals, Littleton, CO, USA) and with the three mouse monoclonal antibodies 3H9, 5C11, and 12H6, as described above, HMGB1 with a rabbit polyclonal anti-high mobility group box-1 (HMGB1) (1:1000, Cat. no. ab18256, Abcam, Cambridge, UK), ERK1/2 with a rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), ICAM-1 with a mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), and VCAM-1 with a mouse monoclonal anti-vascular cell adhesion-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.).
Techniques: Inhibition, Migration, Sterility, Software, Standard Deviation