The chorioallantoic membrane assay as an in vivo model for the study of human bone and tissue engineering

Abstract

Poor vascularization at the fracture site is one of the main limitations of successful bone healing. To improve vascularisation, new tissue engineering approaches aim to design scaffolds that allow not only osteogenesis but also stable capillary invasion: angiogenesis. Importantly, the efficacy and safety of novel bone tissue engineering constructs needs to be tested in vivo before the constructs can be used clinically, which typically involves invasive surgical procedures in murine hosts.
The present thesis proposes the use the chorioallantoic membrane (CAM) from the developing chick as a less sentient in vivo model to explore human bone healing, and the application of this system as a novel screening platform for biomaterials in tissue engineering. To test this model, human bone cylinders were extracted from fresh human femoral heads, stimulated with an injury using a drill-hole defect, and implanted on the CAM or cultured in vitro for a period of 7-9 days. Micro-computed tomography (µCT) was used to quantify the magnitude of bone volume changes together with histological analysis to follow bone repair.
Bone cylinders were vascularised by the CAM blood vessels and remained viable following implantation throughout the culture period. Histological evaluation showed formation of cell condensations in proximity to avian capillaries (immunocytochemistry positive for Sox9 and Runx2). New extracellular matrix (Collagen type II, X) deposition and osteoclast activity (Cathepsin K) on the CAM-implanted cylinders, correlated with a significant increase in bone volume as determined by µCT analysis (p<0.05) and demonstrated the ability of the human-avian system asa test/screen for biomaterials.
In summary, the current studies demonstrate the potential of the CAM assay as a surrogate blood supply for viable human bone tissue. This human-avian system offers a simple refinement model for animal research and a step towards a humanized in vivo model not only for tissue engineering, but for many other disciplines.

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Identifiers

ORCID for Richard Oreffo: orcid.org/0000-0001-5995-6726

ORCID for Nicholas Evans: orcid.org/0000-0002-3255-4388

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