亚砷酸和柔红霉素对NB4细胞Annexin Ⅱ表达及纤溶活性的影响

Influence of arsenic trioxide and daunorubicin on the expression of annexin Ⅱ and fibrinolytic activity in NB4 cells

目的 观察亚砷酸（ATO）和柔红霉素（DNR）对急性早幼粒细胞白血病细胞系NB4细胞膜联蛋白（Annexin,Ann）Ⅱ的表达及其促纤溶活性的影响.方法 用ATO和DNR分别处理NB4细胞24～72 h,用流式细胞术和实时定量PCR方法检测细胞表面Ann Ⅱ的表达及其mRNA的水平;用发色底物法测定其促纤溶活性.结果 与其他白血病细胞系相比,NB4细胞高表达AnnⅡ,NB4、HL-60、K562和A3细胞Ann Ⅱ阳性细胞率分别为（94.5±1.6）%、（40.1±2.1）%、（36.3±1.5）%和（11.8±2.5）%,其促纤溶活性也最强,代表纤溶活性的吸光度（A）值为0.68±0.02.与ATO、DNR及抗Ann Ⅱ单抗作用后NB4细胞促纤溶活性明显下降,分别下降至原来的60.4%、35.8%和26.0%.经ATO和DNR处理48 h均能下调NB4细胞AnnⅡ[Ann Ⅱ阳性细胞率分别为（55.46±4.72）%和（27.00±6.18）%]及其mRNA的表达.结论 ATO和DNR可通过下调NB4细胞Ann Ⅱ蛋白及其mRNA的表达,纠正NB4细胞的高纤溶状态.

Objective To study the expression of annexin Ⅱ （ Ann Ⅱ ） and the fibrinolytic activity in NB4 cells and their alterations in the presence of arsenic trioxide（ATO） and daunorubicin（DNR）. Methods Leukemia cell line NB4 was treated with ATO or DNR for 24 ～ 72 h. Cell surface expression of Ann Ⅱ and its mRNA were analysed by flow cytometry and real time PCR, respectively, the fibrinolytic activity by chromogenic assay. Results Compared with other acute leukemia cell lines, the expression of Ann Ⅱ on untreated NB4 cells was relatively higher. The Ann Ⅱ positive cell rates on NB4, HL-60, K562, and A3 cells were （94.5 ± 1.6） %, （40.1 ± 2.1 ） %, （ 36.3 ± 1.5 ） % and （ 11.8 ± 2.5 ） %, respectively. The fibrinolytic activity of NB4 cells was the greatest with a A value of 0.68 ±0.02. The fibrinolytic activity of NB4 cells was obviously decreased by ATO, DNR or monoclonal antibody against Ann Ⅱ , being decreased by 60.4%,35.8% and 26.0% of the pretreatment level, respectively. The expressions of Ann Ⅱ and its mRNA in NB4 cells were decreased dramatically after ATO and DNR treated for 48 h. Annexin Ⅱ positive cells rate were （55.46 ± 4.72） % and （27.00 ± 6.18） %, respectively. Conclusion NB4 cells have strong ability to enhance the catalytic efficiency of the t-PA-dependent plasminogen activation and Ann Ⅱ on the cell membrane contributes to this activity. Its high fibrinolytic activity can be corrected by ATO and DNR through down-regulating Ann Ⅱ.