title: Sphingomyelinase treatment of low density lipoprotein and cultured cells results in enhanced processing of LDL which can be modulated by sphingomyelin.
abstract: The addition of neutral sphingomyelinase from S. aureus to the medium of rat intestinal epithelial cell cultures (IEC-6) containing added human low density lipoprotein (LDL) resulted in two- to fivefold increases in LDL uptake and degradation. This overall effect was shown to be the combined result of sphingomyelinase activity on the composition of the LDL particle and a separate action directly on the cells when native LDL was incubated with sphingomyelinase from S. aureus followed by removal of the sphingomyelinase. Analysis of sphingomyelinase-treated LDL showed that > 95% of the sphingomyelin (SM) was hydrolyzed, but no changes were observed in all the other components of the LDL particle. This modified LDL particle (SM(-)LDL) was also bound and degraded at higher rates than control LDL in a variety of cell lines, e.g., HepG2, GM-43, and CHO-K1 cells. No evidence of increased aggregation of SM(-)LDL could be observed. The increased processing of SM(-)LDL was due to enhanced affinity to LDL receptors and not to an increase in LDL receptor number. When sphingomyelinase from S. aureus was added to the medium of IEC-6 or GM-43 cells, which were processing SM(-)LDL, further increases in SM(-)LDL processing were observed, which were primarily due to greatly enhanced cellular degradation of SM(-)LDL, with little change in receptor binding and cell association. Since there was little sphingomyelin remaining in SM(-)LDL, it was assumed that the action of sphingomyelinase on the cells resulted in the enhanced degradation. In support of this concept, previous addition of sphingomyelin to cells growing in lipoprotein-deficient medium followed by addition of SM(-)LDL greatly inhibited the degradation of the apolipoprotein of SM(-)LDL. On the other hand, addition of sphingomyelin concomitantly with SM(-)LDL did not inhibit degradation. These results are interpreted to indicate that there may be two pathways for cellular processing of sphingomyelin, one of which may be a determinant in the lysosomal processing of the apolipoprotein of LDL. In support of this concept, addition of desipramine, an inhibitor of lysosomal sphingomyelinase, to IEC-6 cells in culture greatly inhibited the degradation of 125I-labeled LDL without affecting the receptor binding and cell association. Overall, these results suggest that sphingomyelin may play a modulatory role in cellular cholesterol homeostasis by regulating uptake of LDL as well as LDL processing.
authors: Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati, OH 45267-0524.
authors: A K
authors: AK
authors: Gupta
authors: H
authors: H
authors: Rudney
mesh_terms: D000818:Animals; D001693:Biological Transport, Active; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000737:chemistry / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D051381:Rats; D013108:Sphingomyelin Phosphodiesterase / Q000494:pharmacology*; D013109:Sphingomyelins / Q000378:metabolism; D013211:Staphylococcus aureus / Q000201:enzymology
chemical_list: D008077:Lipoproteins, LDL; D013109:Sphingomyelins; D013108:Sphingomyelin Phosphodiesterase