Partial characterization of proteoglycans synthesized by human glomerular epithelial cells in culture

Confluent adult and fetal human glomerular epithelial cells were incubated for 24 h in the presence of [³H]-amino acids and [³⁵S]sulfate. Two heparan-³⁵SO₄ proteoglycans were released into the culture medium. These ³⁵S-labeled proteoglycans eluted as a single peak from anion exchange chromatographic columns, but were separable by gel filtration on Sepharose CL-6B columns. The larger heparan-³⁵SO₄ proteoglycan eluted with the column void volume and at a K_av of 0.26 from Sepharose CL-4B columns. The most abundant medium heparan-³⁵SO₄ proteoglycan was a high buoyant density proteoglycan similar in hydrodynamic size (Sepharose CL-6B K_av 0.23) to those previously described in glomerular basement membranes and isolated glomeruli. Heparan-³⁵SO₄ chains from both proteoglycans were 36 kDa. A smaller proportion of Sepharose CL-6B excluded dermatan-³⁵SO₄ proteoglycan was also synthesized by these cells. The predominant protein cores of both medium heparan-³⁵SO₄ proteoglycans were approximately 230 and 180 kDa. A hybrid chondroitin/dermatan-heparan-³⁵SO₄ proteoglycan with an 80-kDa protein core copurified with the smaller medium heparan-³⁵SO₄ proteoglycan. This ³⁵S-labeled proteoglycan appeared as a diffuse, chondroitinase ABC sensitive 155-kDa fluorographic band in sodium dodecyl sulfate-polyacrylamide gels after the Sepharose CL-6B K_av 0.23 ³⁵S-labeled proteoglycan fraction was digested with heparitinase. The heparitinase generated heparan sulfate proteoglycan protein cores and the 155-kDa hybrid proteoglycan fragment had molecular weights similar to those previously identified in rat glomerular basement membrane and glomeruli using antibodies against a basement membrane tumor proteoglycan precursor (Klein et al. J. Cell Biol. 106, 963-970, 1988). Thus, human glomerular epithelial cells in culture are capable of synthesizing, processing, and releasing heparan sulfate proteoglycans which are similar to those synthesized in vivo and found in the glomerular basement membrane. These proteoglycans may belong to a family of related basement membrane proteoglycans.