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Table 2 Comparison of aortic ring assay applications and novelty

From: Angiogenic potency evaluation of cell therapy candidates by a novel application of the in vitro aortic ring assay

  Nicosia and Ottinetti 1990 [41] Present study 2017
Application/novelty • Developed a quantitative and reproducible angiogenesis assay for cell-free compounds
• Tested the effect of inhibitory/stimulatory soluble factors on angiogenesis
• Modified a quantitative and reproducible angiogenesis assay for coculturing cells
• A novel application to study the angiogenic function and potency of cell therapy candidates for clinical application and banking
Set up • Thoracic rat aorta embedded in fibrin or collagen
• Prepared agarose wells to support aortic rings
• Endothelial media: MCDB 131 to a mixture of 1:1 with DMEM F12 (FBS-free)
• Stimulation of angiogenesis with mouse sarcoma 180
• Studied inhibition of angiogenesis using hydrocortisone
• Thoracic rat aorta embedded between two layers of Matrigel™
• Support of angiogenesis from EHS mouse sarcoma extract (Matrigel™)
• Endothelial basal media supplemented with 2% FBS and 1% penicillin/streptomycin for cocultures
• Prelabeled MSCs cocultured with developing endothelial networks for tracing
Analysis • Daily counting of newly developed microvessels (15 days) using bright-field microscopy
• Florescent imaging of endothelial networks using FVIII-Ra immunohistochemistry
• Quantified radial network growth and network loops at day 5 following MSC coculture
• Live-cell florescence imaging of MSC migration, integration site preference and morphology (coverage) in endothelial networks
• Developed methods to extract MSCs and endothelial cells to perform downstream analysis including flow cytometry and qPCR
  1. FBS fetal bovine serum, MSC mesenchymal stromal cell