title: Isolation and preliminary characterization of proteoglycan aggregates from cultured dermal fibroblasts.
abstract: A1 proteoglycan fractions were prepared by isopycnic cesium chloride density gradient centrifugation from early passage cultured fibroblasts derived from the skin of human infants. In five different fibroblast strains, A1 preparations from the medium and the cells, respectively, contained 36-63% and 59-79% of the sulfated proteoglycans applied to the gradients. A subpopulation of A1 proteoglycans was excluded from Sepharose CL-2B columns under associative conditions (0.5 M guanidium chloride (GdmCl]. The excluded proteoglycans comprised 45-55% of the medium and 28-40% of the cell-associated A1 preparations, showed reversible association-dissociation behaviors on Sepharose CL-2B columns, were composed primarily of chondroitin sulfate and dermatan sulfate, and behaved as monomers with a Kav of 0.33 following mild reduction and alkylation. These data indicate that the excluded peaks were supramolecular aggregates composed of proteoglycan subunits. The mechanism of aggregate formation was examined in A1D1 and D1 fractions. Hyaluronic acid induced aggregation of medium A1D1 proteoglycans in one cell strain, but not in two others. Aggregation was not observed with hyaluronic acid in any cell-associated A1D1 fraction. D1 medium proteoglycans chromatographed on Sepharose CL-2B columns under associative conditions showed two types of aggregates; one was disrupted with 4 M GdmCl indicating that a portion of the aggregate formed through noncovalent interactions that may not require interaction with hyaluronic acid. A second type of aggregate persisted in 4 M GdmCl and after mild reduction and alkylation of the core protein but was disrupted by 0.2% sodium dodecyl sulfate, suggesting that aggregation had occurred through proteoglycan interaction with lipids or hydrophobic proteins. These data indicate that human skin fibroblasts synthesize proteoglycans which share some of the properties of chondrocyte proteoglycans, but also have distinctive macromolecular characteristics.
mesh_terms: D002356:Cartilage / Q000378:metabolism; D002478:Cells, Cultured; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007231:Infant, Newborn; D046911:Macromolecular Substances; D008297:Male; D011509:Proteoglycans / Q000302:isolation & purification* / Q000378:metabolism; D012867:Skin / Q000378:metabolism*
chemical_list: D046911:Macromolecular Substances; D011509:Proteoglycans