[An experimental research of tissue engineered submandibular gland cells growing on collagen sponge scaffold]

To make an experimental research of the tissue engineered rat submandibular glands (SMG) cells growing on a collagen sponge scaffold under an optimal culture condition.

The Wistar rat (8 days old) SMG cells of the second generation were seeded on the surface of the collagen sponge scaffold (5 mm X 5 mm x 2 mm) and were cultured under a physiologically optimal condition for 3 weeks. At 1, 2 and 3 weeks, the cultured cells were observed on their shapes and structures by the histological examination and the scanning electron microscopy. The cultured cells underwent the immunohistochemistry research (the cytokratin 8.13, CK8. 13; alpha-smooth muscular actin, alpha-SMA) staining performed at 3 weeks of the culture, and the amylase activity analysis (the Amano method) performed at 1 day, 1, 2 and 3 weeks of the culture for an evaluation on the secretion function of the cells; the ultrastructures of the cells were also observed by the transmission electron microscopy for an identification of their origins.

The observation under the scanning electron microscope showed that at 1 week after the cell-seeding, the seeded cells were attached to the collagen sponge scaffold surface, with no cell process formed; at 2 weeks the cells increased, with formation of the cell process that was anchored on the collagen sponge scaffold surface; and at 3 weeks, the scaffold surface-attached cells increased, with formation of the filiform fibers in the surface layer of the cells. The immunohistochemistry staining showed that the cultured epithelial cells of SMG were strongly positive for the specific antibody of CK8. 13, and the myoepithelial cells were positive for the specific antibody of alpha-SMA. The transmission electron microscopy showed that in the surface layer of the cultured epithelial cells of SMG the microvilli, plasm crease, and zymogen granules were observed, with a big and oval-shaped nucleus in the cell, and mitochondria and rough endoplasmic reticulum in the cytoplasm of the cell. The amount of amylase secreted by the cells cultured with the collagen sponge scaffolds increased at a different degree with an extension of the culturing time.

The collagen sponge has a satisfactory cell compatibility, and the SMG cells cultured with this kind of collagen sponge can keep their abilities of proliferation and differentiation and their function of secretion. Therefore, this kind of cultured SMG cells can be used as the tissue-engineered cells seeded in the scaffold.