Surface adhesion and attachment factors in bone morphogenetic protein-induced chondrogenesis in vitro.

Bone morphogenetic protein (BMP) induces cartilage and bone development in embryonic and post-fetal life. Bone morphogenetic protein-induced chondrogenesis in outgrowths of muscle connective tissues on various furrowed and unfurrowed fibronectin-coated substrata was observed by biochemical and histologic methods. The substrata consisted of cellulose acetate membranes, atelocollagen, denatured autoclaved demineralized bone matrix, bone matrix deactivated by extraction of BMP with guanidine hydrochloride (GuHCl), and aggregates of insoluble BMP and associated noncollagenous proteins (BMP/NCP) prepared from bovine bone. The furrows, which were mechanically cut into the substrata, were intended to increase surface area and provide extra spaces for cell proliferation under compression. The extent to which extracellular attachments were required for induced cartilage development was reflected in the quantity of cartilage formed when BMP/NCP, either in insoluble or in water-soluble form, was introduced was greatest on GuHCl-extracted bone matrix. On cellulose acetate and atelocollagen, BMP-induced cartilage development was relatively scanty. A substratum of bone matrix, denatured by autoclaving at a minimum of 125 degrees, permitted cell-to-cell adhesion but not cell-to-substratum attachments; the end product was loose fibrous connective tissue only. In contrast, cartilage development occurred on surfaces of undissolved particles of BMP/NCP. Water-soluble human BMP induced development of masses of amorphous cartilage. Even after it was extracted with GuHCl, rat bone matrix may have retained trace amounts of endogenous BMP. Thus, when the requirements of cells for cell-to-cell adhesion and cell-to-substratum attachment, including mechanical factors such as furrows to enlarge surface area, were met, cartilage development was a manifestation of temporal, spatial, and BMP distribution patterns. In situ hybridization and immunofluorescent microscopy with the aid of antirecombinant BMP antibiotics may provide new information about morphogenesis.