Blastogenic Factor from Unstimulated Leukocyte Cultures: Further Evidence for Heterogeneity, and Mechanism of Production

Abstract Blastogenic factor derived from unstimulated leukocyte cultures was studied. The present studies have shown that the properties of the blastogenic factor which stimulates only allogeneic cells (allogeneic blastogenic factor) are different in some respects from those of the blastogenic factor which stimulated autologous cells as much as allogeneic cells (autologous blastogenic factor). Activity of the allogeneic blastogenic factor was labile to exposure to temperature of 60°C, and was concentrated in the precipitate obtained by ultracentrifugation of cell-free culture medium (CFM) at 100,000 × G. In contrast, activity of the autologous blastogenic factor was resistant to the same temperature, and could not be sedimented by ultracentrifugation. The kinetics of in vitro production of the allogeneic blastogenic factor was also different from those of the autologous blastogenic factor. While activity of the autologous blastogenic factor was almost constant throughout the incubation period, that of the allogeneic blastogenic factor reached a maximum after 4 to 5 days of incubation and thereafter decreased. These findings suggest that there may be separate molecules responsible for the allogeneic and autologous blastogenic activities. The blastogenic factor prepared from mixed cultures of intact cells and treated allogeneic cells was also studied. Although the difference was not significant (p = 0.09) when leukocytes were treated with actinomycin D, the CFM was more active for cells from the donor of the treated cells than for cells from the donor of the intact cells in eight out of 12 experiments. The ultracentrifuged precipitate (probably allogeneic blastogenic factor) was significantly more stimulatory toward cells from the donor of the treated cells than from the donor of the intact cells. When leukocytes were treated with Escherichia coli endotoxin, both the CFM and the ultracentrifuged precipitate stimulated cells from the donor of the treated cells more than cells from the donor of the intact cells. Mytomycin C did not have any effect on the blastogenic activity of the CFM and the ultracentrifuged precipitate prepared from these cultures.