甲异靛对JAK2/V617F杂合突变细胞系SET2细胞凋亡和增殖作用及其机制的研究  

The impact of meisoindigo on apoptosis and proliferation of SET2 cell line by JAK-STAT pathway

作　　者：吕成兰 刘晋琴[1] 陈猛 陈兵 肖志坚[1] Lyu Chenglan;Liu Jinqin;Chen Meng;Chen Bin;Xiao Zhijian(Institute of Hematology and Blood Disease,Chinese Academy of Medical Sciences and Peking Union Medical College,Tianjin 300020,China;The Affiated Hospital of Nanjing University Medical School,Nanjing Drum Tower Hospital,Nanjing 210009,China)

机构地区：[1]中国医学科学院、北京协和医学院血液病医院(血液学研究所),实验血液学国家重点实验室,天津300020 [2]南京大学医学院附属鼓楼医院,210009

出　　处：《中华血液学杂志》2019年第1期29-34,共6页Chinese Journal of Hematology

基　　金：实验血液学国家重点实验室自主课题(Z17-03);中国医学科学院医学与健康科技创新工程(2016-I2M-1-001);协和学者与创新团队发展计划.

摘　　要：目的观察甲异靛对JAK2/V617F杂合突变细胞系SET2细胞凋亡及增殖的影响,并探讨JAK-STAT信号通路在其中所起的作用。方法不同浓度甲异靛(0、5、10 μmol/L)作用于SET2细胞24、48、72 h后,采用流式细胞术检测细胞凋亡。不同浓度甲异靛(0、5、10、20 μmol/L)作用于SET2细胞24、48、72、96 h后,CCK8法检测细胞增殖。不同浓度甲异靛(0、5、10、20 μmol/L)作用于SET2细胞12 h后,在甲基纤维素半固体培养基进行集落培养,Western blot法测定JAK-STAT信号通路相关蛋白及凋亡相关蛋白表达水平。结果在0、5、10 μmol/L甲异靛作用于SET2细胞后24、48和72 h,细胞凋亡率均随药物浓度的增大而增高(P<0.01)。0、5、10、20 μmol/L甲异靛作用SET2细胞后,随着药物浓度增加,细胞增殖受抑制(P<0.01)。不同浓度甲异靛处理SET2细胞后,其集落形成率随药物浓度增加而降低[0 μmol/L甲异靛组为(4.48±1.19)%,20 μmol/L甲异靛组为(2.55±0.36)%,Dunnett P=0.020]。不同浓度甲异靛作用于SET2细胞12 h后,JAK2、P-JAK2、P-STAT1、P-STAT3、P-STAT3和STAT5的表达水平均有随药物浓度增加而逐渐下调的趋势,抗凋亡蛋白BCL-2、BCL-XL的表达水平随药物浓度增加而下调,促凋亡蛋白BID、BIM的表达水平随药物浓度增加而上调。结论甲异靛对Ph阴性骨髓增殖性肿瘤细胞系SET2细胞具有明显的促进凋亡、抑制增殖作用,伴随着抗凋亡蛋白表达下调及促凋亡蛋白表达上调,其作用机制可能与甲异靛抑制JAK-STAT信号通路有关。Objective To observe the effect of meisoindigo on apoptosis and proliferation of JAK2/V617F heterozygous mutation cell line-SET2 cell line to further explore the role of JAK-STAT pathway in this effect.Methods Cell apoptosis after treated with different concentration of meisoindigo (0,5,and 10 μmol/L) was evaluated by flow cytometry at different time points (24,48,72 h).Cell proliferation with CCK8 test was evaluated at different time points (24,48,72,96 h) after administered with different concentration of meisoindigo (0,5,10,and 20 μmol/L).After treatment with different concentration of meisoindigo (0,5,10,and 20 μmol/L),SET2 cells were collected after 12 h,and then cultured in incomplete methylcellulose-based medium for clone formation.JAK-STAT signaling pathway and apoptosis related protein by Western blot test were evaluated 12 h after administered with different concentration of meisoindigo (0,5,10,and 20 μmol/L).Results At different time points after treated with meisoindigo,the apoptosis rate of SET2 cell lines increased (P<0.01) with the inhibited proliferation (P<0.01),and the decreased clone formation rate of SET2 cell lines [0 μmol/L meisoindigo:(4.48±1.19)%,20 μmol/L meisoindigo:(2.55±0.36)%;Dunnett P=0.020] in the presence of augmented concentrations of meisoindigo.At 12 hours after administration with meisoindigo,the reduced expressions of JAK2,P-JAK2,P-STAT1,P-STAT3,P-STAT3,STAT5,the decreased anti-apoptosis proteins BCL-2,BCL-XL and the increased pro-apoptosis protein BID,BIM were observed in the presence of increased concentrations of meisoindigo.Conclusion Meisoindigo played an important role during the apoptosis and the inhibition of proliferation in ph-negative myeloproliferative neoplasm cell-SET2 cell lines,which might be related to the inhibition of JAK-STAT signaling pathway with up-regulation of pro-apoptosis protein and down-regulation of anti-apoptosis protein.

关 键 词：甲异靛 JAK-STAT信号通路 SET2细胞 

分 类 号：R733.7[医药卫生—肿瘤]