Journal: Arteriosclerosis, Thrombosis, and Vascular Biology
Article Title: Matrix-Binding, N-Cadherin–Targeting Chimeric Peptide Inhibits Intimal Thickening but Not Endothelial Repair in Balloon-Injured Carotid Arteries
doi: 10.1161/ATVBAHA.123.319400
Figure Lengend Snippet: N-cadherin–targeting chimeric peptide inhibited migration of human smooth muscle cells (SMCs), but not endothelial cells (ECs), and did not impact viability. Scratch-wounded human aortic SMCs (HASMCs; A ) or human umbilical vein endothelial cells (HUVECs; C ) were treated with N-cadherin–targeting chimeric peptide or scrambled control peptide and imaged after 48 hours. The area of the wound gap was subtracted from and divided by the initial gap area to give a percentage of the wound that had closed. For viability measurements, HASMCs ( B ) or HUVECs ( D ) were treated with N-cadherin–targeting chimeric peptide or scrambled control peptide and allowed to proliferate for 24 hours. Metabolic activity was assessed by colorimetric change of water-soluble tetrazolium salt, measured as absorbance at 450 nm, with background at 610 nm subtracted. Apoptosis was measured in HUVECs by staining for cleaved caspase 3 (CC3) and counting positively stained cells ( E ). Permeability was measured in HUVECs grown on transwell membranes with 0.4-μm pores where the top chamber media replaced with 5 μM fluorescein isothiocyanate (FITC)–dextran and N-cadherin peptide or scrambled control. Media in the abluminal ( bottom ) chamber was sampled into a 96-well dish and luminescence at 530 nm read by plate reader. FITC-dextran concentration calculated against a standard curve ( F ). Data shown are means±SEM; n≥3 biological replicates. Statistical significance was determined by 2-way ANOVA with Tukey multiple comparisons ( A–D ) or Mann-Whitney U tests ( E and F ).
Article Snippet: Human umbilical vein ECs (HUVECs) and human arterial SMCs (HASMCs) were purchased from Lonza (Basel, Switzerland) and cultured in EGM-2 (endothelial growth medium-2; Lonza) or SmGm-2 (smooth muscle growth medium-2; Lonza) media supplemented with the included bullet kit according to the manufacturer’s instructions.
Techniques: Migration, Activity Assay, Staining, Permeability, Concentration Assay, MANN-WHITNEY
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