[Expression and significance of melanoma antigen gene-3 in endoplasmic reticulum stress-induced apoptosis of K562 cells].
This study was aimed to explore the expression and significance of melanoma antigen gene-3 (MAGE-3) in endoplasmic reticulum stress-induced apoptosis. After the treatment of leukemia cell line K562 and its multidrug-resistant cell line K562/A02 by thapsigargin, intracellular calcium concentrations ([Ca(2+)]i) in K562 and K562/A02 were measured by fluorescence spectrophotometer with calcium sensitive fluorescence indicator Fura-2/AM; expression changes of glucose-regulated protein 78 (GRP78) were detected by Western blot; morphological change of apoptotic cell was investigated by AO/EB fluorescent staining under fluorescent microscope; apoptosis rate was determined by terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP nick end labelling (TUNEL) staining; the expression of MAGE-3 gene mRNA was detected by RT-PCR. The results showed that (1) thapsigargin induced the enhancement of [Ca(2+)]i with different extent in K562 and K562/A02 cells, and the enhancement of [Ca(2+)]i was dose-dependent in experiment range. At the same time, thapsigargin induced upregulation of GRP78 protein expression and typical apoptotic changes of K562 and K562/A02 cells, apoptotic rate was also dose-dependent in experiment range. The [Ca(2+)]i in K562/A02 cells were higher than that in K562 cells. (2) in the course of endoplasmic reticulum stress-induced apoptosis by thapsigargin, the expression of MAGE-3 gene mRNA was remarkably downregulated. Moreover, the expression of MAGE-3 gene mRNA in K562/A02 cells was higher than that in K562 cells. It is concluded that (1) thapsigargin induces endoplasmic reticulum stress-induced apoptosis of K562 and K562/A02 cells in experiment range, and this may be associated with downregulation of MAGE-3 mRNA expression or MAGE-3 gene may participates in the regulation of endoplasmic reticulum stress-induced apoptosis. (2) MAGE-3 gene may possess anti-apoptotic activity, multidrug resistance in K562/A02 cells can be associated with [Ca(2+)]i increase and upregulation of MAGE-3 expression.
Feng
Xian-Qi
XQ
Department of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China. fxqnsm2000@yahoo.com.cn
Xiao
Juan
J
Nie
Shu-Min
SM
Liu
Fang
F
You
Yong
Y
Zou
Ping
P
D000951:Antigens, Neoplasm / Q000235:genetics*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling; D020014:K562 Cells; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones / Q000378:metabolism; D009363:Neoplasm Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019284:Thapsigargin / Q000494:pharmacology
D000951:Antigens, Neoplasm; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; C072013:MAGEA3 protein, human; D018832:Molecular Chaperones; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D019284:Thapsigargin; D002118:Calcium