Characterisation of chondroitin/dermatan sulphate proteoglycans synthesised by rat mammary myoepithelial and fibroblastic cell lines

Abstract

Chondroitin sulphate proteoglycans were isolated from the culture medium of rat mammary gland fibroblast (Ram 27) and myoepithelial (Rama 401) cell lines which had been labelled with [ 35S]sulphate. Chromatography on Sepharose CL-4B indicated that the Rama 401 proteoglycan was larger than the Rama 27 proteoglycan ( Kav values 0.47 and 0.56, respectively). Treatment of the proteoglycans with alkaline NaBH 4 yielded chondroitin sulphate chains with average M rvalues of 37 000 (Rama 401) and 21 000 (Rama 27). Structural analysis of the glycosaminoglycan chains indicated that both were co-polymers of chondroitin and dermatan sulphate although there were differences in the amounts and distribution of the disaccharide repeating units. The M rvalues of the core proteins, determined by immunoblotting, were about 43 000 and 46 000 (Rama 27) and 44 500 (Rama 401). Using an antibody to chondroitin sulphate proteoglycan in immunofluorescence experiments, the proteoglycan was demonstrated on the surface of both cell lines. Rama 27 cells additionally possessed an extensive fibrous extracellular matrix which also stained with the antibody. Staining of sections of lactating mammary gland suggested that the proteoglycan was present in the basement membrane as well as the stromal connective tissue. The presence of chondroitin sulphate proteoglycan in the basement membrane was confirmed by ultrastructural immunolocalisation.