Chorioallantoic membrane

The chorioallantoic membrane is composed of three layers. The first is the chorionic epithelium that is the external layer present immediately below the shell membrane.[2] It consist of epithelial cells that arise from chorionic ectoderm. The second is the intermediate mesodermal layer that consists of mesenchymal tissue formed by the fusion of the mesodermal layer of the chorion and the mesodermal layer of the allantois. This layer is highly vascularized and rich in stromal components. The third is the allantoic epithelium that consists of epithelial cells arising from the allantoic ectoderm. It forms a part of the wall of the allantoic sac.

Both the epithelial layers are separated from the mesodermal layer by basement membranes.[3]

The Chorioallantoic membrane performs the following functions:

The CAM functions as the site of gaseous exchange for oxygen and carbon dioxide between the growing embryo and the environment. Blood capillaries and sinuses are found in the intermediate mesodermal layer allows close contact (within 0.2 μm) with air found in pores of the shell membrane of the egg.[4]

The chorionic epithelial layer contains the calcium transporting region of the CAM, and thus is responsible for the transport of calcium ions from the egg shell into the embryo for the purpose of ossification of the bones of the developing embryo.[2][5] The CAM also helps in maintaining the acid-base homeostasis in the embryo.[6] Finally the allantoic epithelium serves a barrier to the allantoic cavity, and acts in a selectively permeable manner by permitting the absorption of water and electrolytes, as well as maintains a barrier against the toxins and waste materials stored inside the allantoic cavity.[2]

The development of the CAM is similar to that of the allantois in mammals. Its growth starts from day 3 of embryonic development. Development of the allantois occurs extra embryonically from the ventral wall of the endodermal hindgut. Partial fusion of the chorion and allantois occurs between days 5 and 6. By day 10, there is an extensive formation of capillary network. The complete differentiation of the CAM is complete by day 13.[7][5]

Chorioallantoic membranes can be cultivated either outside (ex-ovo) or inside of the shell (in-ovo).

CAM provides several features such as ease of access, and the rapid development of the membrane structure, presence of an immunodeficient environment,[10] ease of visualization for imaging techniques ranging from microscopic to PET scans.[7] Thus, it makes for a suitable model for a number of research applications in the field of biological and biomedical research:

• Vascular development and angiogenesis.
• Xenograft studies.[10]
• Study of tumour growth and differentiation.[11]
• Wound repair studies.
• Toxicology studies.
• Nanomaterials assessments.[12]
• Drug delivery.
• Study of molecules affecting with angiogenic and anti-angiogenic activities.
• Culturing of viruses like Herpes Simplex Virus, etc.[13][14]
• Drug screening studies.
• Radiotherapy related studies.
• Allergenicity and toxicity studies.
• Helminth cultivation.[15]
• Oncological investigations.[16]

The advantages of using CAM are:

•   It is easy to use as compared to other animal models.
•  Assays can be visualized real time using very simple to highly complex visualizing techniques.[7]
•  Rapid vascular growth.
•  Cost effective, easy to access.
•  The circulatory system is completely accessible making the delivery of intravenous molecules easy.
•  Assays take relatively less time.
•  Easily reproducible and reliable.

Despite the numerous advantages, there are a number of disadvantages associated with the use CAMs:

• Sensitivity to modifications in environmental conditions.[5]
• Limited availability of reagents like antibodies due to avian origin.
• Non-specific inflammatory reaction after 15 days of development.[5]
• Difficulty of distinguishing the formation of new capillaries from the already existing vascular network.[7]
• Differences in metabolism of drugs as compared to mammals.