An in vitro system to study listericidal capacity of macrophages from separate mice: resident macrophages exhibit different activation patterns

Abstract

An in vitro system with macrophages from individual mice was established to study their listericidal capacity. Because no antibiotics were used, bacterial killing was really due to macrophages in short-term culture. To restrict the extracellular growth of bacteria, cell culture medium was changed at 1-h intervals. We demonstrated that intracellular growth of listeria in macrophage pools from untreated animals varies considerably. Obviously, preactivated macrophages are constantly present, so that the common procedure of using macrophage pools from several animals is no longer acceptable. In addition, we demonstrated that in vitro mixtures of listeria-immune macrophages of one animal with cells from untreated animals at different ratios exhibit enhanced bacterial killing above a mere additive effect. Consequently, by using macrophages from individual untreated mice, we found that cells of different animals exhibited various activation stages, although unstimulated, inbred specific-pathogen-free mice of the same age, weight, and sex were used. When equal numbers of macrophages from untreated separate animals were mixed in vitro, intracellular growth of listeria was only moderate; that is, the number of preactivated macrophages of the individual animals determined listerial growth in the pooled preparation. Furthermore, we showed that identical doses of phorbol myristate acetate exerted different effects on the listericidal activities of macrophages as a function of their preactivation states. These experiments clearly demonstrate the advantage of using macrophages from individual mice for in vitro studies of macrophage activation.