Proteoglycan synthesis by skeletal muscle undergoing bone matrix-directed transformation into cartilage in vitro.
Myoblasts and fibroblasts of embryonic skeletal muscle reproducibly form chondrocytes when cultured on demineralized bone in vitro. The transformation occurs in 3 morphologically defined phases, with disappearance of the myoblast phenotype preceding the appearance of fibroblast-like cells and finally chondrocytes. Proteoglycan synthesis in these cultures was investigated by labeling at prechondrogenic (5 days) and postchondrogenic (6-12 days) stages with (35S)sulfate and [6-3H]glucosamine. Labeled material elutes from associative Sepharose CL-2B columns as two major included peaks, which correspond to proteoglycan monomer and a material of lower molecular size. Control cultures, cultured upon gels of type I collagen, fail to synthesize monomer-like material and contain solely a material of lower molecular size. Demineralized bone-derived monomer was rechromatographed under dissociative conditions in an attempt to detect the presence of small aggregates. Again, only a single peak of sulfate and glucosamine-labeled material appears. The data further show that the monomer resembles that of embryonic cartilage in glycosaminoglycan chain size (Mr = 8.6-12.2 X 10(3] and composition (mainly chondroitin 4-sulfate). Aggregated monomer forms a shoulder of the monomeric peak and comprises only 5% of the sulfated material. Fifteen to thirty-four per cent of the monomer elutes as aggregate after addition of rooster comb hyaluronic acid (HA). Failure to aggregate appears to be related to endogenous synthesis of short chain HA. Synthesis of long chain HA may constitute a rate-limiting step in chondrogenesis. Material of lower molecular size, from cultures grown on demineralized bone, bind to exogenous HA, whereas the elution pattern of sulfated material from control cultures remains essentially unchanged. These latter data suggest that proteoglycans of low hydrodynamic size may participate in the early formation of proteoglycan aggregate.
D000818:Animals; D001857:Bone Matrix / Q000502:physiology*; D002356:Cartilage / Q000254:growth & development*; D002850:Chromatography, Gel; D005260:Female; D066298:In Vitro Techniques; D024510:Muscle Development; D009132:Muscles / Q000378:metabolism; D011247:Pregnancy; D011509:Proteoglycans / Q000096:biosynthesis*; D051381:Rats; D013997:Time Factors
D011509:Proteoglycans