摘要
本研究探讨三氧化二砷(arsenic trioxide,ATO)诱导NB4、MR2细胞凋亡过程中蛋白磷酸酶2A(PP2A)活性与表达的变化。不同浓度ATO单用或与冈田酸(OKA)联合作用于NB4、MR2细胞,然后采用MTT法测定药物对细胞增殖的影响,用瑞氏染色法观察细胞形态学变化,流式细胞术测定细胞凋亡率,丝/苏氨酸磷酸化酶检测试剂盒分析药物对细胞内PP2A活性的影响,Western blot检测细胞内PP2A各亚基表达。结果表明:蛋白磷酸酶抑制剂OKA能明显增强ATO对NB4、MR2细胞的增殖抑制;OKA能促进ATO诱导的NB4、MR2细胞凋亡;ATO诱导NB4、MR2细胞凋亡过程中PP2A活性均下降,且随ATO浓度的增加PP2A活性下降越明显;与OKA连用后PP2A活性下降更加明显;在ATO诱导NB4、MR2细胞凋亡过程中,PP2A/A表达较对照组均明显减少,而B和C亚基表达与对照组比较变化不明显。结论:在ATO诱导的NB4、MR2细胞凋亡过程中,细胞内PP2A/A表达减少,PP2A活性降低,且随ATO浓度增加PP2A活性下降越明显,抑制PP2A活性有助于ATO诱导的NB4、MR2细胞凋亡。
This study was aimed to investigate the change of expression and activity of protein phosphatases type 2A （PP2A） during the apoptosis of NB4 and MR2 cells induced by Arsenic trioxide（ATO）. NB4 and MR2 cells were incubated with Okadaic acid（ OKA ） （0.5 nmol/L）, ATO （0.5 - 2.0 μmol/L）, and the combination of OKA and ATO at the same doses as in the single-agent treatment respectively. Then the proliferation of NB4 and MR2 cells was determined by MTT assay, the morphologic changes of cells were evaluated by Wright＇s staining, the apoptosis rates were detected by flow cytometry. At last, the activities of PP2A were evaluated by the serine/threonine phosphatase assay system, and the levels of PP2A subunits were detected by Western blot analysis. The results showed that ATO inhibited proliferation of NB4 and MR2 cells, and the inhibition rates of ATO on the two cells significantly increased after the addition of OKA. OKA could augment the apoptosis of NB4 and MR2 cells induced by ATO. During the apoptosis of NB4 and MR2 cells, the activity of PP2A decreased with increasing concentration of ATO, and OKA augmented the inhibitory effect of ATO on the activity. The level of PP2A structural subunit （PP2A-A） decreased during ATO-induced apoptosis of NB4 and MR2 cells, that expressions of B and C subunits of PP2A were relatively unaltered. It is concluded that the activity of PP2A decreases with increasing concentration of ATO during the apoptosis of NB4 and MR2 cells, and the decrease of the activity of PP2A maybe is related to the repression of expression of PP2A -A subunit; the inhibition of the activity of PP2A can promote the ATO induced apoptosis of NB4 and MRL cells.
出处
《中国实验血液学杂志》
CAS
CSCD
2008年第5期1021-1025,共5页
Journal of Experimental Hematology
