TURNOVER OF ENDOGENOUS CERAMIDE IN CULTURED NORMAL AND FARBER-FIBROBLASTS

De novo synthesis and turnover of endogenous ceramide in cultured skin fibroblasts from patients affected with Farber lipogranulomatosis wer e studied by biosynthetical labeling of cellular sphingolipids with [C -14]serine. The cellular uptake of [C-14]serine and incorporation into de novo synthesized ceramide was similar in normal and Farber fibrobl asts, with a half life of newly synthesized ceramide of 2.7 h in norma l and diseased cells. Newly synthesized ceramide was found to be chann eled directly into biosynthesis of complex sphingolipids rather than c ontributing to the pool of accumulated ceramide in Farber fibroblasts. The degradation of ceramide generated by the catabolism of complex sp hingolipids in Farber cells was greatly delayed compared with control fibroblasts, with differences in the amount of radiolabeled cellular c eramide becoming evident after 6 h chase time. Individual Farber cell lines differed from each other in the amount of accumulated ceramide; however, no correlation was found between ceramide accumulation and re sidual acid ceramidase activity as determined in vitro. In addition, t he amount of radiolabeled sphingomyelin was significantly increased in Farber fibroblasts suggesting a delayed degradation of this compound in this ceramide storage disorder. We propose biosynthetical labeling of endogenous ceramide with [C-14]serine, in addition to other establi shed methods, as a highly sensitive and reliable method for the diagno sis of Farber disease, allowing semiquantitative measurement of cerami de accumulation in cultured skin fibroblasts of patients affected with Farber lipogranulomatosis.