Soluble factors secreted by macrophage-like cells in vitro cause osteoprogenitor cell detachment

Explant cultures of adult rabbit bone were obtained from a variety of skeletal sites, with the intention of using the cells in a new type of bone graft. The explant cultures contained large numbers of esterase-positive, alkaline phosphatase-negative, TRAP-negative macrophage-like cells (MP), as well as osteoblast-like osteoprogenitor (OP) cells (alkaline phosphatase-positive). OP numbers were assessed by cell counts and MTT assay. The presence of the MP cells appeared to give rise to a reduction in OP numbers in culture, through cell detachment and cell lysis. In addition, after passaging, many OP were unable to reattach to the culture vessels in the presence of MP, depending upon the surface area available for reattachment. The presence of tumor necrosis factor alpha (TNFalpha) in culture medium from these cell cultures was demonstrated by a specific enzyme-linked immunosorbent assay (ELISA). A direct relationship was demonstrated between MP numbers and TNFalpha concentration and an inverse relationship between MP numbers and OP numbers in co-cultures. This was also found when OP cells were exposed to different concentrations of rTNFalpha, in place of the MP. Incubation with anti-TNFalpha inhibited the effect of MP and TNFalpha on OP. These results suggest that MP are able to cause detachment and lysis of OP cells, probably by secretion of a soluble factor in vitro which may be TNFalpha.