Temporal relationships between ceramide production, caspase activation and mitochondrial dysfunction in cell lines with varying sensitivity to anti-Fas-induced apoptosis

被引:47
作者
Rodriguez-Lafrasse, C [1 ]
Alphonse, G [1 ]
Broquet, P [1 ]
Aloy, MT [1 ]
Louisot, P [1 ]
Rousson, R [1 ]
机构
[1] Lyon Sud Med Sch, INSERM, U189, F-69921 Oullins, France
关键词
Jurkat cells; SCC61; cells; sphingolipid metabolism; SQ20B cells;
D O I
10.1042/0264-6021:3570407
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
To clarify the chronology of events leading to anti-Fas-induced apoptosis, and the mechanisms of resistance to this death effector, we compared the response kinetics of three tumour cell lines that display varying sensitivity to anti-Fas (based on levels of apoptosis), in terms of ceramide release, mitochondrial function and the caspase-activation pathway. In the highly sensitive Jurkat cell line, early caspase-8 activation, observed from 2 h after treatment, was chronologically associated with an acute depletion of glutathione and the cleavage of caspase-3 and poly-ADP ribosyl polymerase (PARP), followed by a progressive fall in the mitochondrial transmembrane potential (Delta psi (m)), between 4 and 48 h after treatment. Ceramide levels began to increase 2 h after the addition of anti-Fas (with no increase during the first hour), and increased continuously to 640% of control cells at 48 h. In the moderately sensitive SCC61 adherent cells, comparable results were observed, though with lower levels of ceramide and a delay in the response kinetics, with apoptotic cells becoming flotant. Finally, despite early cleavage of caspase-8 at 2 h, and a sustained level of activation until 48 h, no apoptotic response was observed in anti-Fas-resistant SQ20B cells. This was confirmed by a lack of ceramide generation and mitochondrial changes, and by the absence of any detectable cleavage of caspase-3 or PARP. Inhibition of caspase processing, and amplification of endogenous ceramide signalling by pharmacological agents, allowed us to establish the order of cellular events, locating ceramide release after caspase-8 activation and before caspase-3 activation, and demonstrating a direct involvement for ceramide release in mitochondrial dysfunction. Furthermore, these experiments provide strong arguments for the role of endogenous ceramide as a key executor of apoptosis, rather than as a consequence of membrane alterations.
引用
收藏
页码:407 / 416
页数:10
相关论文
共 47 条
[1]   EFFECT OF TRICYCLIC ANTI-DEPRESSANTS ON SPHINGOMYELINASE AND OTHER SPHINGOLIPID HYDROLASES IN C6 CULTURED GLIOMA-CELLS [J].
ALBOUZ, S ;
VANIER, MT ;
HAUW, JJ ;
LESAUX, F ;
BOUTRY, JM ;
BAUMANN, N .
NEUROSCIENCE LETTERS, 1983, 36 (03) :311-315
[2]  
Bezombes C, 2001, FASEB J, V15, P297
[3]   Common regulation of apoptosis signaling induced by CD95 and the DNA-damaging stimuli etoposide and γ-radiation downstream from caspase-8 activation [J].
Boesen-de Cock, JGR ;
Tepper, AD ;
de Vries, E ;
van Blitterswijk, WJ ;
Borst, J .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1999, 274 (20) :14255-14261
[4]   CD95 (Fas/APO-1) induces ceramide formation and apoptosis in the absence of a functional acid sphingomyelinase [J].
Boesen-de Cock, JGR ;
Tepper, AD ;
de Vries, E ;
van Blitterswijk, WJ ;
Borst, J .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1998, 273 (13) :7560-7565
[5]   Involvement of MACH, a novel MORT1/FADD-interacting protease, in Fas/APO-1- and TNF receptor-induced cell death [J].
Boldin, MP ;
Goncharov, TM ;
Goltsev, YV ;
Wallach, D .
CELL, 1996, 85 (06) :803-815
[6]   Apoptosis without caspases: an inefficient molecular guillotine? [J].
Borner, C ;
Monney, L .
CELL DEATH AND DIFFERENTIATION, 1999, 6 (06) :497-507
[7]   Caspases induce cytochrome c release from mitochondria by activating cytosolic factors [J].
Bossy-Wetzel, E ;
Green, DR .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1999, 274 (25) :17484-17490
[8]   Fas/CD95/Apo-I activates the acidic sphingomyelinase via caspases [J].
Brenner, B ;
Ferlinz, K ;
Grassmé, H ;
Weller, M ;
Koppenhoefer, U ;
Dichgans, J ;
Sandhoff, K ;
Lang, F ;
Gulbins, E .
CELL DEATH AND DIFFERENTIATION, 1998, 5 (01) :29-37
[9]   Fas-mediated apoptosis is modulated by intracellular glutathione in human T cells [J].
Chiba, T ;
Takahashi, S ;
Sato, N ;
Ishii, S ;
Kikuchi, K .
EUROPEAN JOURNAL OF IMMUNOLOGY, 1996, 26 (05) :1164-1169
[10]  
Chmura SJ, 1997, CANCER RES, V57, P1270