Abstract
Objective To investigate apoptosis in mouse leukemia cell(WEHI-3) induced by Thapsigargin and its mechanism.Methods Apoptosis induced by Thapsigargin was examined by flow cytometry and Agarose gel electrophoresis of DNA and TUNEL staining.Free calcium(i)was determined by Fura-2 fluorescein load techinique.RT-PCR was used to analyze the mRNA of Caspase-12.Results After exposure to Thapsigargin of 0.5,1 and 2 μmol/L,WEHI-3 cells were induced to apoptosis.The apoptotic index was examined by Annexin-V/PI double-stained,Agarose gel electrophoresis of DNA and TUNEL.The results showed that the apoptotic index of the experimental groups was significantly higher than that of the control group(For example:the apoptotic index of the experimental groups determined by Annexin-V/PI double-stained were(25.3±3.2)%,(46.7±3.9)% and(70.2±2.3)%,respectively,which were significantly higher than the apoptotic index of the control group which was(7.6±0.4)%(F=26.52,P0.01);and the apoptotic index was in dose dependent manner.After exposure to Thapsigargin,that i in the experimental group was significantly higher than that of the control group[(156.5±10.3)nmol/L,(180.3±15.6) nmol/L and(10.7±15.3)nmol/L vs.(78.3±11.2)nmol/L(F=21.26,P0.01)].The mRNA expression of Caspase-12 increased with an increase in Thapsigargin concentration.Conclusion Thapsigargin can induce the WEHI-3 to apoptosis by Endoplasmic reticulum stress of i overload,Ca2+ may play a promising target for cancer therapy.
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