SULFATIDE AND SPHINGOMYELIN LOADING OF LIVING CELLS AS TOOLS FOR THE STUDY OF CERAMIDE TURNOVER BY LYSOSOMAL CERAMIDASE - IMPLICATIONS FOR THE DIAGNOSIS OF FARBER-DISEASE

The ceramide turnover by lysosomal ceramidase in intact, living cells was investigated by loading radiolabeled sulfatide or sphingomyelin in situ on skin fibroblasts and lymphoid cells. The cells originated fro m normal individuals and from patients with acid ceramidase deficiency (Farber disease). While fibroblasts from individuals with Farber dise ase exhibited some impairment in the degradation of the ceramide produ ced by sulfatide hydrolysis, lymphoid cells from individuals with Farb er disease metabolized the ceramide as readily as did normal cells, su ggesting the existence in lymphoid cells of a nonlysosomal degradation pathway for the sulfatide-derived ceramide, In contrast, sphingomyeli n loading in the presence of serum showed a considerably decreased tur nover of ceramide in both fibroblasts and lymphoid cells from individu als with Farber disease. Further methodologic variation led to the use of LDL-associated radioactive sphingomyelin; LDL-association promoted the targeting of exogenous sphingomyelin to lysosomes. As a result, a n almost complete deficiency of ceramide degradation was found in cell s from severely affected patients with Farber disease. Our data with t his novel method show that sphingomyelin loading of intact living cell s is a simple, alternative means for determining ceramide degradation by lysosomal ceramidase and for diagnosing Farber disease. (C) 1995 Ac ademic Press, Inc.