Partial isolation and characterization of a chemotactic factor from adult bovine bone for mesenchymal cells

Demineralized adult bone matrix has the capacity to initiate de novo ectopic endochondral bone formation 2-3 weeks following intramuscular implantation into suitable hosts. An early step in this process is the migration of mesenchymal cells to the implant site; these cells later differentiate into cartilage and bone. Adult bone has been shown to contain a number of bioactive factors, such as chemotactic factors for various cell types, including osteoblasts. We have used embryonic chick limb bud mesenchymal cells to construct an in vitro assay for testing chemotactic activity derived from bone matrix extracts. With a modified Boyden chamber, water-soluble components from a 4 M guanidinium chloride extract of demineralized adult bovine bone matrix were found to stimulate the directional migration of these chick embryonic limb bud mesenchymal cells as well as embryonic muscle-derived fibroblasts and cells derived from embryonic skin. The chemotactic activity was destroyed by treatment with heat (100 degrees C) or trypsin. Partial purification by molecular sieve chromatography suggested that the chemotactic factor(s) has a molecular weight of between 50,000 and 90,000. This factor can be separated from bone matrix-derived chondrogenic stimulating activity by either ion exchange or molecular sieve chromatography. These observations confirm that bone matrix contains a chemoattractant for mesenchymal cells that may be important for in vivo recruitment of cells as part of the process of ectopic bone formation or in cases of bone repair.