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Cell-density Dependent Changes of Glycosphingolipid Biosynthesis in Cultured Human Skin Fibroblasts

An involvement of glycosphingolipids (GSL) in regulation of cell growth and contact inhibition of growth at saturated cell density has been documented in several nontransformed cell lines. In this study, the glycosphingolipid biosynthesis was investigated in the sparse and the confluent cell populations (CP) of cultured human skin fibroblasts (HSF). The HSF-populations were metabolically pulse labelled with 14C-galactose (24-48h). The amounts of 14C-radioactivity (cpm) incorporated into extracted and purified total cellular GSL- fractions were counted by ß-scintillation and the individual GSL-species were HPTLC-separated and visualized by autoradiography. The relative labelling (%) of individual newly synthesized GSL- species was detected by densitometric scanning of autoradiographic GSL-patterns. The incorporation of 14C-label into total GSL- fractions per cell increased significantly as the cell-density increased. The total newly synthesized GSL-pattern (100%) of sparse HSF-CP showed a significant predominance of the ganglioside-species (GG ; 70%) over the neutral GSL-species (NGSL ; 30%), with GM2 as the main species followed by CMH. The relative labelling of all GG-species moderately decreased while the NGSL-species increased as the cell density increased. Oppositely, within the total newly synthesized GSL-pattern of confluent HSF-CP (contact-inhibited), the NGSL-species (67%) predominated over the GG-species (33%). The most abundant species were by turns CMH, CTH and GM3 while GM2 was approx. 10-fold lower than in the sparse HSF-CP. The results support the hypothesis that GSL expressed at the cell surface might be involved in cell-density dependent regulation of cell growth.

glycosphingolipids ; cultured human skin fibroblasts ; cell density

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