ED2+ macrophages increase selectively myoblast proliferation in muscle cultures

We have previously shown by coculturing myoblasts and macrophages that myotube formation is strongly increased in vitro by the presence of an acid stable, heat-labile, soluble growth factor(s) secreted by macrophages. In this paper we obtained macrophages from peritoneal washing which also contained limited amounts of other cells such as lymphocytes and mesothelial cells. We here demonstrate that an ED2-positive (ED2+) macrophage subpopulation is responsible for myoblast enhanced proliferation. ED2+ macrophages were separated by a magnetic-activated cell sorter (MACS) using a monoclonal antibody against ED2, a membrane antigen peculiar to macrophages. Both ED2+ macrophages and their conditioned medium increased myotube formation when added to primary muscle cultures. Furthermore we demonstrate that muscle growth induced by macrophages is mainly the consequence of an increased myoblast proliferation by showing the presence of an increased number of MyoD-positive (MyoD+) myonuclei.