We used a double-labeling technique to compare the metabolism of complex carbohy- drates by skin fibroblasts from patients with rheumatic heart disease and from age- and sex-matched normal controls. Fibroblasts were subcultured through a similar number of passages prior to their growth in the presence of "C- or3H-labeled glucosamine, a precursor of complex carbohydrates. At the preconfluent, confluent, and postconfluent stages of growth, the culture medium and trypsin digest of the3H- oruC-labeled fibroblasts were combined, incubated with Protease, then successively fraction- ated on Biogel P-2, Biogel P-100, and DE-52 cellulose. Individual fractions were assayed for radioactiv- ity, and the radioactive elution profiles of labeled complex carbohydrates were compared. These profiles showed that trypsin digestion releases quantitatively more high molecular weight complex carbohydrates and proportionately less heterogeneous glycopeptides from normal fibroblasts, as compared to fibroblasts from rheumatic patients. Chemical analysis of the components of these complex carbohydrates revealed that complex carbohydrates of fibroblasts from rheumatic patients contained more glycoproteins and proportionately less hyaluronic acid than complex carbohydrates of normal fibroblasts. In addition, normal fibroblasts secreted significantly more high molecular weight complex carbohydrates into the medium than fibroblasts from rheumatic patients. These differences were confined to the medium recovered from fibroblast cultures grown to post confluency but not to earlier stages of growth. These findings suggest the possible presence of a biochemical alteration in the synthesis of complex carbohydrates by fibroblasts of rheumatic individuals. The basis and exact nature of this alteration remain to be defined.Circ Res 46: 651-659, 1980