This study investigates perineural invasion (PNI) in metastasizing cancers using a chicken egg model. The dorsal root ganglia (DRG) from mice are grafted onto the chorioallantoic membrane (CAM) to observe cancer cell behavior towards nerve tissues.
In certain metastasizing cancers, tumor cells extend away from the site of origin and penetrate the nerve tissues, a phenomenon called perineural invasion or PNI.
PNI can be demonstrated in ovo on the surface of the chorioallantoic membrane or CAM, which mimics the basement membrane of the human epithelial tissue.
To begin, take a viable, fertilized chicken egg with its shell partially opened, exposing a fully developed CAM.
Next, isolate the dorsal root ganglia or DRG from a mouse’s spinal cord. DRG is a cluster of sensory neurons using which the cancer cells invade the perineural space.
Incubate the DRG in a suitable medium with a fluorescent dye to label them for easy visualization. Now, transfer the DRG onto the CAM surface without rupturing the membrane.
Cover the egg with a transparent film to prevent microbial contamination. Thereafter, incubate the egg to facilitate DRG integration into CAM.
Following incubation, seed the fluorescently labeled cancer cells on a dry surface of CAM, away from the DRG. The dry surface minimizes the spread of cancer cells and allows regular tumor formation.
Seal the eggshell opening and incubate it to allow DRG-cancer cell interaction. Under the microscope, the cancer cells can be observed growing towards the engrafted DRG, demonstrating PNI.
For grafting of a dorsal root ganglion onto the chorioallantoic membrane, use fine, sterile forceps to carefully grasp one ganglion in the culture dish and gently wash the tissue in a container of fresh HBSS. Place the ganglion onto the chorioallantoic membrane of one egg, being careful not to puncture the membrane, and cover all of the egg openings with sterile transparent film dressings. When all of the eggs have been grafted, return them to the humidifying egg incubator for 2 days without rotation.
At the end of the incubation, transfer the eggs to the laminar flow cabinet, and use scissors and forceps to open the transparent film dressing. Next, add 0.5 to 1 million fluorescently labeled tumor cells in 5 microliters of HBSS onto the chorioallantoic membrane of each egg, about 2 millimeters from each dorsal root ganglion, keeping a uniform distance between the ganglia and the cells. Then, cover the eggs with a new film dressing and return the eggs to the egg incubator for 7 days without rotation.
繁體中文
