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目的探讨葛根提取物(PL)对慢性粒细胞白血病(CML)细胞株K562的凋亡诱导作用及其可能的分子机制。方法以不同浓度(0、2.5、5、10和20 mg/ml)PL处理K562细胞,24 h后光镜下观察形态学改变,MTT法测定细胞增殖抑制,Hoechest 33258荧光染色及FITC-AnnexinⅤ/PI双染法检测凋亡率变化,半定量RT-PCR及Western blot方法检测Bcr/abl、Bcl-2、p53、Fas/FasL蛋白表达。结果PL对K562细胞有明显的增殖抑制作用,并观察到典型的细胞凋亡形态变化,细胞凋亡率与浓度呈正相关;不同浓度PL干预后Bcr/abl基因在mRNA和蛋白水平呈浓度依赖性下调,而Bcl-2基因则无明显变化;p53表达呈浓度依赖性上调;Fas/FasL表达无明显变化。结论不同浓度PL能有效诱导K562细胞凋亡;其机制可能通过在mRNA和蛋白水平下调Bcr/abl基因,上调p53基因表达而实现。
Objective To investigate the apoptosis-inducing effect of Pueraria lobata (PL) on chronic myeloid leukemia (CML) cell line K562 and its possible molecular mechanism. Methods K562 cells were treated with various concentrations of PL (0, 2.5, 5, 10, 20 and 20 mg / ml) for 24 h. Morphological changes were observed under light microscope. Cell proliferation inhibition was measured by MTT assay. Hoechst 33258 fluorescence staining and FITC-Annexin V / PI double staining was used to detect the changes of apoptosis rate. The expression of Bcr / abl, Bcl-2, p53, Fas / FasL protein was detected by semi-quantitative RT-PCR and Western blot. Results The proliferation of K562 cells was inhibited by PL, and the morphological changes of typical apoptotic cells were observed. The apoptotic rate was positively correlated with the concentration of Bcr / abl mRNA and protein Down-regulated, while the Bcl-2 gene had no significant change. The expression of p53 was up-regulated in a concentration-dependent manner. There was no significant change in the expression of Fas / FasL. Conclusion Different concentrations of PL can effectively induce apoptosis of K562 cells. The mechanism may be through down-regulating Bcr / abl mRNA and protein levels and up-regulating p53 gene expression.