[Mechanism of Fas/FasL signal transduction pathway in K562 cell apoptosis induced by diallyl disulfide].
The aim of this study was to investigate the effect of diallyl disulfide (DADS) on the apoptosis of K562 cells and to explore the mechanism of K562 apoptosis induced by DADS. The K562 cells were treated with different concentrations of DADS for 24, 48 and 72 hours. The concentrations of DADS were as follows: 0 (control group), 10, 20, 40 and 80 mg/L. The morphologic changes of leukemia K562 cells treated with DADS were observed by Hoechst33 258 staining. The apoptosis of K562 cells treated with different concentrations of DADS for 24, 48 and 72 hours was analyzed by flow cytometry. The mRNA expression changes of Fas and FasL were detected by reverse transcription-polymerase chain reaction (RT-PCR) after K562 cells were treated with different concentrations of DADS for 48 hours. The results indicated that the characteristics of apoptosis in K562 cells induced by DADS were as follows: reduction of nucleus, chromatin condensation and nuclear membrane rupture. The flow cytometry with PI straining showed that after 24 hours of DADS treatment the apoptosis rate of K562 cells increased from 11.60 ± 0.83% at the concentration of 10 mg/L to 37.94 ± 0.87% at the concentration of 40 mg/L. The apoptosis rate of K562 cells increased from 37.94 ± 0.87% (24 hours) to 47.02 ± 0.66% (72 hours) after treatment with DADS of 10 mg/L increasing to 40 mg/L DADS. The Fas mRNA expression levels of the related apoptotic genes increased after K562 cells were treated with different concentrations of DADS for 48 hours, while FasL mRNA expression decreased significantly after DADS treatment for 48 hours, compared with those in the control group (p < 0.05). It is concluded that DADS can induce the apoptosis of human leukemia K562 cells in a time-and concentration-dependent manners. The activation of Fas/FasL pathway may play an important role in the K562 cell apoptosis induced by DADS, which is associated with increasing Fas gene expression and decreasing FasL gene expression.
Available from: Pritha Dasgupta
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ABSTRACT: Diallyl disulfide (DADS), the major organosulfur component of processed garlic is very effective in chemoprevention of several types of cancers; however, its detailed mechanism is yet to be divulged. Present study shows antiproliferative activity of DADS against human leukemic cell-lines, mainly U937. DADS induced transient G2/M phase arrest, which is evident from FACS analysis. The results revealed that a significant transcriptional induction of p21 happened in early hours of treatment, which is due to increased nuclear translocation of NF-κB and its specific binding to p21 promoter. However, in the later hours, G2/M arrest is lost leading to apoptosis via intrinsic mitochondria-mediated pathway through generation of reactive oxygen species followed by changes in mitochondrial membrane potential. Western blots indicate release of cytochrome-c, activation of caspase-3, cleavage of PARP1, and finally decrease in bcl-2 levels. In addition, inactivation of NF-κB by its inhibitor BAY 11-7085 causes early onset of apoptosis without any transient G2/M arrest. Thus, in conclusion, DADS induces reversible G2/M arrest through NF-κB mediated pathway in human leukemic cell lines, like U937, K562, and Jurkat, lacking wild type p53. However, G2/M arrest is lost owing to the incapability of the damage repair system that leads to apoptosis.
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