Isolation of human tumor cells that produce cementum proteins in culture

Abstract

We have cultured cells from explants of a human cementum tumor. The cells obtained were multipolar, they formed network-like structures and they were alkaline-phosphatase positive. Immunostaining and Western blots using specific antibodies revealed that these cells produced bone sialoprotein and collagen types I and V, and they also mineralized in vitro. Conditioned medium was mitogenic to fibroblasts and mitogens present were separated by heparin-affinity chromatography. Based on affinity to heparin and antibody-inhibition studies, the heparin fractions were shown to contain cementum-derived growth factor, platelet-derived growth factor and fibroblast growth factors. The cementum tumor cells, but not gingival fibroblasts, were stained positively by an antibody to cementum-derived attachment protein. The attachment protein was separated by immunoaffinity chromatography, and Western blots revealed that the preparation contained 56-kDa and 43-kDa proteins as major bands. Cells pulse-labeled with radioactive amino acids contained a 43-kDa protein as the major component; however, this protein was absent after a cold chase in the presence of cycloheximide, but 56-kDa, 39-kDa and 26-kDa species became prominent. These data indicated that the 56-kDa cementum attachment protein is derived from a 43-kDa precursor. Our data show that the cells cultured from the cementum tumor represent cementum cells capable of synthesizing and secreting cementum proteins in culture.