Slow rise of Ca2+ and slow release of reactive oxygen species are two cross-talked events important in tumour necrosis factor-alpha-mediated apoptosis.
Tumour necrosis factor-alpha (TNF-alpha) was found to be a cell cycle-independent apoptogenic cytokine in cultured fibroblast L929 cells. This assertion is based on the observations (1) TNF-alpha increased the number of cells with hypo-diploid DNA in a time dependent manner as revealed by flow cytometry, and (2) TNF-alpha induced DNA fragmentation as resolved by agarose gel electrophoresis. When cells were exposed to TNF-alpha (50 ng/ml), a slow rise in intracellular free Ca2+ level and a delayed increase in the production of reactive oxygen species (ROS) (both observed 3 h after the addition of TNF-alpha) were observed in fluo-3 and fura-red or dichlorofluorescein loaded cells, respectively. Interestingly, challenge of cells with TNF-alpha in the presence of BAPTA/AM, an intracellular Ca2+ chelator, decreased the release of ROS. Removal of ROS by 4-hydroxy 2,2,6,6-tetra-methyl-piperidinooxy (4OH-TEMPO) blocked the TNF-alpha-mediated Ca2+ rise. Moreover, when cells were exposed to TNF-alpha with both 4OH-TEMPO and BAPTA/AM, more viable cells were found than from treatment with either BAPTA/AM or 4OH-TEMPO. These results suggest that ROS and cellular Ca2+ are two cross-talk messengers important in TNF-alpha-mediated apoptosis.
Ko
S
S
Department of Biochemistry, The Chinese University of Hong Kong, Shatin, NT.
Kwok
T T
TT
Fung
K P
KP
Choy
Y M
YM
Lee
C Y
CY
Kong
S K
SK
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D003497:Cyclic N-Oxides / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D051379:Mice; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D013113:Spin Labels; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D000975:Antioxidants; D002614:Chelating Agents; D003497:Cyclic N-Oxides; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D013113:Spin Labels; D014409:Tumor Necrosis Factor-alpha; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D002118:Calcium; C001803:tempol