TRYPSIN PROMOTES C6 GLIOMA CELL-PROLIFERATION IN SERUM-FREE AND GROWTH FACTOR-FREE MEDIUM

C6 glioma cells could be successively subcultured and maintained in se rum- and growth factor-free medium (SF/GFF medium). C6 cell proliferat ion in SF/GFF medium was positively correlated with the initial cell d ensity at plating. This correlation disappeared when the medium had be en renewed early after cell adhesion (3 h after plating), suggesting t hat C6 cell growth depends on some diffusible factor in the medium bef ore renewal, and that this factor is not secreted from C6 cells in the assay culture but is transferred from the cell suspension. The supern atant of trypsinized C6 cell suspension (SCS), trypsin-EDTA solution f or routine cell harvesting use, and modified trypsin of protein sequen cing grade all promoted C6 cell proliferation at, appropriate dilution s or concentrations under SF/GFF conditions. The growth promoting effe cts of SCS and trypsin-EDTA solution were completely inhibited by soyb ean trypsin inhibitor. These results demonstrate that the serine prote ase trypsin has a proliferative effect on C6 cells continuously subcul tured in SF/GFF medium. In addition, it is suggested that trypsin used for cell dispersion is transferred from cell suspension into the cult ure, where it promotes C6 cell growth after passage in our SF/GFF subc ulture system.