Study on the expression of IkappaB-alpha protein in TNF-alpha induced apoptosis of U937 cells.
OBJECTIVE
To investigate the TNF-alpha induced apoptosis of U937 cells, the expression, degradation and subcellular localization of IkappaB-alpha, and its degradation mechanism.


METHODS
Changes and subcellular loca-lization of IkappaB-alpha were observed by fluorescence microscopy, expression and degradation of IkappaB-alpha protein with N-tosyl-L-phenylalanylchloromethyl ketone (TPCK protease inhibitor) blocking test and apoptosis of U937 cell by flow cytometry.


RESULTS
(1) immunolfluorescence assay showed that IkappaB-alpha localized in cytoplasm only. (2) The level of IkappaB-alpha protein was downregulated after TNF-alpha stimulation, flow cytometry also confirmed the downregulation. (3) The downregulation of IkappaB-alpha protein levels in TNF-alpha induced apoptosis was partially inhibited by TPCK. (4) The apoptosis rate of U937 cells induced by TNF-alpha was (60.73 +/- 1.61)%.


CONCLUSIONS
(1) Degradation of IkappaB-alpha protein during TNF-alpha induced apoptosis of U937 cells suggested the activation of NF-kappaB. (2) TPCK sensitive protease plays an important role in the degradation of IkappaB-alpha protein. (3) TPCK sensitive protease also involved in the apoptosis of U937 cells induced by TNF-alpha.
Chen
Weihua
W
Department of Pediatrics, Tongji Hospital, Tongji Medical College of Huazhong University of Science and Technology, Wuhan 430030, China.
Peng
Guangjie
G
Tang
Aiping
A
Wang
Kaiyan
K
Zhou
Muxiang
M
Wang
Lingyi
L
D017209:Apoptosis / Q000187:drug effects; D015536:Down-Regulation; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D020298:U937 Cells
D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha