lysophosphatidylethanolamine and edelfosine

We have isolated two mutant strains from the murine, macrophage-like cell line, RAW 264.7, that are resistant to the cytotoxic effects of the antineoplastic, platelet activating factor analogue, 1-octadecyl-2-methyl-rac-glycero-3-phosphocholine (ET-18-OMe). The mutants were isolated using a single round of selection to ensure that resistance was due to a single gene defect. These mutants, RAW.R1 and RAW.R23, are approximately 20 times more resistant to ET-18-OMe (ID50 = 15-17 microM) than the parent strain (ID50 = 0.7-1.0 microM). Resistance to ET-18-OMe was due to a 90-95% reduction in the ability to take up and accumulate this compound. The uptake of other choline glycerophospholipids (e.g., platelet activating factor and 1-acyl-2-lyso-sn-glycero-3-phosphocholine) was also severely affected. This defect was not limited to choline glycerophospholipids; the uptake of an ethanolamine glycerophospholipid (1-alkyl-2-lyso-sn-glycero-3-phosphoethanolamine) was reduced by 80%. The uptake of palmitic acid, an amphipathic molecule bearing no phosphate-containing head group, was unaffected in the mutants. There was little metabolism of ET-18-OMe by either the wild-type or mutant cells. Binding of ET-18-OMe appeared to be normal in the mutants, but internalization of pre-bound ET-18-OMe was reduced. Uptake of non-lipid ligands such as horseradish peroxidase, lucifer yellow, and transferrin was normal in the mutants demonstrating that fluid-phase and receptor-mediated endocytosis is functional. The ability to generate mutants displaying a lesion that affects the uptake of both choline and ethanolamine phospholipids demonstrates that these species are internalized by RAW cells through one common primary route or through pathways that share a common factor. These mutants, and this approach to their isolation, offer a system with which to study and define the mechanisms of glycerophospholipid uptake into macrophages as well as other cell types.

Topics: Animals; Cell Line; Drug Resistance; Endocytosis; Glycerophosphates; Glycerylphosphorylcholine; Lysophosphatidylcholines; Lysophospholipids; Macrophages; Mice; Mutation; Phosphatidic Acids; Phospholipid Ethers