[Mitochondrial transmembrane potential loss caused by reactive oxygen species plays a major role in sodium selenite-induced apoptosis in NB4 cells].
OBJECTIVE
To investigate the role of reactive oxygen species (ROS) and ROS-caused mitochondrial transmembrane potential loss in sodium selenite-induced apoptosis in NB4 cells.


METHODS
ROS production was measured by ROS-specific probe DCFH-DA. Sodium selenite mitochondrial transmembrane potential loss was evaluated by flow cytometry with Rh123 staining. Protein levels of cytochrome C, Bid, Bcl-xl, and Bax were measured by Western blot using protein-specific antibodies. NB4 cells were pre-incubated by MnTmPy or BSO before selenite treatment to further confirm the effects of ROS on NB4 cells.


RESULTS
20 micromol/L sodium selenite induced ROS production and mitochondrial transmembrane potential loss in NB4 cells time-dependently. Cytochrome C accumulated in cytoplasm after selenite treatment. Sodium selenite also downregulated Bcl-xl and activated Bax and Bid at protein level. Pretreatment with antioxidant MnTmPy almost fully abrogated the proapoptotic effect of sodium selenite prevented the cleavage of Bid protein and in turn the mitochondrail transmembrane potential loss. On the contrary, pretreatment with BSO intensified the mitochondrail transmembrane potential loss induced by sodium selenite.


CONCLUSIONS
Sodium selenite may induce apoptosis by inducing ROS production in NB4 cells, which leads to the downregulation of Bcl-xl, upregulation of Bax, and cleavage and activation of Bid. Bax and tBid then agregate on mitochondrial membrane, which in turn causes a decrease of mitochondrial transmembrane potential and release of cytochrome C into cytoplasm.
Wei
Wei
W
National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
Han
Bing-she
BS
Guan
Li-ying
LY
Huang
Fang
F
Feng
Lei
L
Yang
Yang
Y
Xu
Cai-min
CM
D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein / Q000096:biosynthesis; D045744:Cell Line, Tumor; D045304:Cytochromes c / Q000378:metabolism; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D018038:Sodium Selenite / Q000494:pharmacology*; D051028:bcl-2-Associated X Protein / Q000096:biosynthesis; D051020:bcl-X Protein / Q000096:biosynthesis
C494826:BAX protein, human; C494811:BCL2L1 protein, human; D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; D017382:Reactive Oxygen Species; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D045304:Cytochromes c; D018038:Sodium Selenite