Induction of apoptosis in human mitogen-activated peripheral blood T-lymphocytes by the ether phospholipid ET-18-OCH3: involvement of the Fas receptor/ligand system.
1. Activated T-cells constitute a target for treatment of autoimmune diseases. We have found that the antitumour ether phospholipid 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3; edelfosine) induced dose- and time-dependent apoptosis in human mitogen-activated peripheral blood T-lymphocytes, but not in resting T-cells. T-lymphocytes were stimulated with phytohemagglutinin and interleukin-2 or with concanavalin A. Apoptosis was assessed by DNA fragmentation through cell cycle and TUNEL analyses, as well as through visualization of internucleosomal DNA fragmentation in agarose gels. 2. The ET-18-OCH3-mediated apoptotic response in activated T-lymphocytes was less intense than in human leukaemic T cell lines, such as Jurkat cells and Peer cells; namely about 25% apoptosis in activated T-cells versus about 46-61% apoptosis in T leukaemic cells after 24 h treatment with 10 microM ET-18-OCH3. 3. The ET-18-OCH3 thioether analogue BM 41.440 (ilmofosine) showed a similar apoptotic capacity to that found with ET-18-OCH3 in activated T-cells, whereas the phospholipid analogue hexadecylphosphocholine (miltefosine) failed to promote this response. 4. The uptake of [3H]-ET-18-OCH3 was much larger in activated T-cells than in resting lymphocytes. 5. Using a cytofluorimetric approach we have found that ET-18-OCH3 induced disruption of the mitochondrial transmembrane potential and production of reactive oxygen species in activated T-cells, but not in resting lymphocytes. 6. ET-18-OCH3 induced an increase in Fas (APO-1/CD95) ligand mRNA expression in activated T-cells, and incubation with a blocking anti-Fas (APO-1/CD95) antibody partially inhibited the ET-18-OCH3-induced apoptosis of activated T-lymphocytes. 7. These results demonstrate that mitogen-activated T-cells, unlike resting lymphocytes, are able to take up significant amounts of ET-18-OCH3, and are susceptible to undergo apoptosis by the ether lipid via, in part, the Fas (APO-1/CD95) receptor/ligand system. This ET-18-OCH3 apoptotic action can be of importance in the therapeutic action of this ether lipid in certain autoimmune diseases.
Cabaner
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C
Centro de Investigación del Cáncer, Instituto de Biología Molecular y Celular del Cáncer, CSIC-Universidad de Salamanca, Spain.
Gajate
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C
Macho
A
A
Muñoz
E
E
Modolell
M
M
Mollinedo
F
F
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D053222:Fas Ligand Protein; D006801:Humans; D020287:In Situ Nick-End Labeling; D008213:Lymphocyte Activation; D008562:Membrane Glycoproteins / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects; D008934:Mitogens / Q000494:pharmacology; D010742:Phospholipid Ethers / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects* / Q000502:physiology; D019014:fas Receptor / Q000502:physiology*
D000970:Antineoplastic Agents; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D008934:Mitogens; D010742:Phospholipid Ethers; D017382:Reactive Oxygen Species; D019014:fas Receptor; C026659:edelfosine; D003513:Cycloheximide