Indomethacin induces apoptosis of K562 cells through activation of caspases and elevation of intracellular free calcium
10.3760/j:issn:0253-2727.2001.05.005
- VernacularTitle:吲哚美辛诱导K562细胞凋亡过程中半胱天冬酶及细胞内游离钙浓度的变化
- Author:
Guangbiao ZHOU
1
;
Guangsen ZHANG
Author Information
1. The Second Affiliated Hospital of Hunan Medical University
- From:
Chinese Journal of Hematology
2001;22(5):241-244
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the indomethacin (IN) induced apoptosis of K562 cells: ① the expression, activation and subcellular localization of caspase-3 and -8;②the change of intracellular free calcium concentration (［fCa2+］i) and its mechanism; and ③ whether the cell apoptosis is cyclooxygenase (cox) dependent or not. Methods Changes and subcellular localization of caspase-3 and -8 were observed by laser scanning confocal microscopy (LSCM); expression and activation of caspase-3 and -8 proteins by Western blotting; changes of intracellular ［fCa2+］i by LSCM coupling with a calcium-fluorescence probe and calcium chelator EGTA blocking test; and cox inhibitor effect by MTT assay. Results ①LSCM assay and Western blotting showed that caspase-3 and -8 localized in cytoplasm and nucleus dispersedly and spottedly and were upregulated with the increasing of IN doses. Western blotting also showed the cleavage and activation of caspase-3 and -8 during IN-induced apoptosis. ②The increase of ［fCa2+］i in K562 cells was parallel to the increase of IN concentration regardless of the presence or absence of EGTA. But with EGTA treatment,［fCa2+］i was much less than that without EGTA treatment. ③Low dose of IN or other cox inhibitors could not exert cytotoxic effects on K562 cells whereas high dose of IN could. Conclusion ①The upregulation and activation of caspase-3 and -8 play a fundamental role in apoptosis induced by IN in K562 cells. Both cytoplasm and nucleus are locations where caspase-3 and -8 localized. ②The increase of ［fCa2+］i may be critical in the modulation of apoptosis; the extracellular calcium influx is the main source of the elevation of ［fCa2+］i in K562 cells and can be blocked by the calcium chelator EGTA;the release of calcium from intracellular calcium store is also an important source of the intracellular calcium which can trigger apoptosis without the extracellular calcium influx.③Apoptosis of K562 cells induced by IN is cox-independent.
