|
pubmed:abstractText |
Ceramide acts as an intracellular second messenger in cellular signal transduction. We examined the effects of two cell-permeable ceramides, C2-ceramide and C6-ceramide, on human monocyte functions. After monocytes were primed with lipopolysaccharide (LPS) or interferon-gamma (IFN-gamma) for 18 hr in suspension culture, they produced a high amount of superoxide (O2-) when triggered by phorbol myristate acetate. C2- or C6-ceramide inhibited O2- release from monocytes primed with LPS (1 ng/ml) or IFN-gamma (100 U/ml), but did not affect unprimed monocytes. An analogue, C2-dihydroceramide, was inactive. C2-ceramide was most effective at 6 microM, and C6-ceramide at 60 microM. C2- or C6-ceramide at these concentrations was not toxic for monocytes, as assessed by trypan blue exclusion and by the 3-[4, 5-dimethylthiazol-2-y1]-2,5 diphenyl tetrazolium bromide (MTT) assay which measures the ability of live cells to produce formazan. C2-ceramide (20 microM) had no effect on the killing of leukaemic cells (HL-60 and K562 cells) by monocytes treated with IFN-gamma, LPS, or both for 18 hr, with killing assessed by an 111 Indium-releasing assay. C2-ceramide (20 microM) induced secretion of low amounts of tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) from the monocytes. But C2-ceramide did not alter the higher secretion of TNF-alpha or IL-1 beta from monocytes treated with IFN-gamma or LPS. Thus the cell-permeable ceramides acted like antagonists of LPS, rather than analogues of LPS, as has been proposed. The results here showed that the signal transduction pathway for O2- release by monocytes differed from that for the cytolysis of leukaemic cells, and confirmed that oxygen radicals are not involved in cytolysis.
|
