Anti-apoptotic effects of probucol are associated with downregulation of Daxx expression in THP-1 macrophage.
OBJECTIVE
To study the relationship between Daxx expression and the antiapoptotic effects of probucol in THP-1 macrophage.


METHODS
Apoptosis of THP-1 derived macrophages was induced by exposure to oxidized low density lipoprotein (oxLDL). The development of apoptosis was determined by flow cytometry analysis and nucleic acid-binding dye acridin orange. Reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting and indirect immunofluorescence were used to evaluate the expression of Daxx and caspase-3 at both mRNA and protein level.


RESULTS
As expected, THP-1 macrophages exposed to 100 mg/l oxLDL for 48 h exhibited typical morphologic changes of apoptosis, including condensed chromatin and shrunken nucleus. oxLDL treatment markedly increased Daxx expression in a time- and dose-dependent manner, and facilitated Daxx translocation from cytoplasm to nucleus. The percentage of cells with Daxx in nuclei was significantly increased from 8 to 59%. Treatment with probucol (50 micromol/l) for 4 h prior to exposure to oxLDL significantly inhibited Daxx expression and THP-1 macrophage apoptosis by 61.3%. Furthermore, oxLDL enhanced caspase-3 expression with increased mRNA and protein levels, but without obvious change in translocation of caspase-3 (the cells with nuclear Daxx: 14 vs 8%). In contrast, probucol attenuated oxLDL-stimulated caspase-3 expression in THP-1 macrophages.


CONCLUSIONS
OxLDL-induced apoptosis of THP-1 macrophage is associated with Daxx up-regulation; while inhibition of apoptosis by probucol is related to decreased Daxx expression and nuclear translocation.
Su
Bo
B
Division of Pharmacoproteomics, Institute of Pharmacy & Pharmacology, Nanhua University, Changsheng Western Road 28#, Hengyang, 421001, Hunan, China.
Yang
Yun-Bo
YB
Tuo
Qin-Hui
QH
Zhu
Bing-Yang
BY
Lei
Xiao-Yong
XY
Yin
Weidong
W
Liao
Duan-Fang
DF
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics* / Q000378:metabolism; D000924:Anticholesteremic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D056970:Co-Repressor Proteins; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008077:Lipoproteins, LDL / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018832:Molecular Chaperones; D009000:Monocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D011341:Probucol / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors
D048868:Adaptor Proteins, Signal Transducing; D000924:Anticholesteremic Agents; D056970:Co-Repressor Proteins; C106985:DAXX protein, human; D008077:Lipoproteins, LDL; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D012333:RNA, Messenger; C099252:oxidized low density lipoprotein; D053148:Caspase 3; D011341:Probucol