三七皂甙对造血细胞GATA-1和GATA-2转录调控蛋白的诱导作用  

Up-regulation of transcription factors GATA-1 and GATA-2 induced by panax notoginosides in hematopoietic cells

作　　者：高瑞兰 徐卫红 林筱洁 陈小红 吴超群[2] 

机构地区：[1]浙江中医学院附属医院,杭州310006 [2]复旦大学生命科学院

出　　处：《中华血液学杂志》2004年第5期281-284,共4页Chinese Journal of Hematology

基　　金：国家自然科学基金资助项目 ( 3 0 0 70 93 3 )

摘　　要：目的　观察三七总皂甙 (PNS)对锌指结构GATA族转录调控蛋白的诱导作用 ,探讨PNS在造血细胞内的信号传递途径。方法　选择恰当浓度的PNS作用于正常人骨髓单个核细胞、HL 6 0、K5 6 2、CHRF 2 88和Meg 0 1细胞 ,2周后提取核蛋白 ,用GATA 1和GATA 2抗体行Western印迹杂交 ,检测GATA 1和GATA 2的表达情况。用3 2 P标记的GATA双链寡核苷酸探针行电泳带移动阻滞试验 (EMSA)和抗体胶移动实验 ,检测GATA DNA复合物及亚型。结果　PNS能够促进人骨髓红系、粒系祖细胞增殖。Western印迹杂交显示PNS诱导K5 6 2、CHRF 2 88和Meg 0 1细胞的GA TA 1和GATA 2蛋白表达量增高 ,分别是未经处理细胞的 1 .5～ 2 .8倍和 2 .0～ 3.1倍 ;EMSA结果表明PNS诱导三株细胞的GATA DNA结合复合物条带密度明显增高 ;HL 6 0细胞在PNS处理前后均未显示GATA活性。抗体胶移动实验证明PNS诱导的GATA DNA复合物的主要成分为GATA 1和GATA 2。免疫沉淀显示PNS诱导的GATA 1和GATA 2蛋白均处于磷酸化的功能激活状态。结论　PNS可通过诱导GATA 1和GATA 2蛋白合成增加 ,与相关基因上游调控区的启动子和 (或 )增强子结合的活性增高 ,而调控与造血细胞增殖。Objective To observe the role of panax notoginosides (PNS) in up regulation of GATA family transcription factors, and explore intracellular signal pathway of PNS in the proliferation of hematopoietic cells. Methods Human bone marrow cells were incubated with different concentrations of PNS for colony forming assay. Human cell lines HL 60, K562, CHRF 288 and Meg 01 were incubated with PNS (10 mg/L) for 14 days. The cell nuclear proteins were extracted and analyzed by Western blot with antibodies against GATA 1, GATA 2. Electrophoretic mobility shift assay (EMSA) and antibody gel supershift assay was performed using 32 P labeled GATA consensus oligonucleotide which contains binding site for GATA transcription factors. Results PNS could promote the proliferation of CFU GM and CFU E and induce the expression of GATA 1, GATA 2 . The nuclear proteins of both GATA 1 and GATA 2 in K562, CHRF 288 and Meg 01 cells treated by PNS were increased by (1.5～2.8) and (2.0～3.1) fold over untreated cells respectively. GATA binding activity initiated by PNS was apparently elevated to form higher density band of GATA DNA complex. While there was no detectable change in HL 60 cells before and after PNS treatment. The predominant GATA binding complex was mainly attributable to both GATA 1 and GATA 2 proteins being in phosphorylated status. Conclusion PNS can induce the synthesis of transcription factors GATA 1 and GATA 2 and enhance their DNA binding activity, which could play a role in the up regulation of the expression genes related to proliferation and differentiation in hematopoietic cells.

关 键 词：三七皂甙 造血细胞 PNS GATA-1 GATA-2 调控蛋白 诱导作用 转录调控 

分 类 号：R285[医药卫生—中药学]