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Table 3 Aortic ring assay

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

Advantages Limitations
• Cost-effective because the aorta is waste tissue from endpoint animal studies
• From one aorta, a high yield of replicates can be obtained (approximately 20 rings per adult animal)
• Rapid set up including aorta isolation and embedding
• Evaluates key steps of angiogenesis including matrix degradation, cell migration, proliferation and morphogenesis into tubular endothelial network
• In addition to endothelial cells, includes cell types important for angiogenesis such as resident pericytes and fibroblasts
• Endothelial cells have not been preselected by passaging and thus are in a quiescent state at starting point of the assay, reflecting in vivo conditions
• Quiescent endothelial cells respond by proliferating and differentiating into tubular networks
• Evaluates properties of cocultured candidate cell types including the ability to respond to signals of aortic tissue, induce migration, ECM processing and homing to endothelial networks
• Cell-to-cell connections can be observed using fluorescence microscopy
• Net effects on angiogenesis can be quantified using image analysis software to assess various network properties (network radial growth, loops, branches and nodes)
• Vessel outgrowths occur from a major vessel while in vivo angiogenesis occurs typically from micro vessels
• Takes 3–5 days for initial endothelial network to develop
• Variability in angiogenic response can also occur between animal’s due to strain, age and gender
• Lack of blood flow (limitation shared with other in vitro and ex vivo angiogenesis assays)
• Angiogenic vessel growth is in three dimensions, rendering imaging and quantification difficult
• Outgrowth vessels regress over time (2 weeks), thereby limiting long-term analysis
  1. ECM extracellular matrix