摘要
为了进一步研究三七总皂苷 (PNS)对转录调控蛋白AP 1家族的主要成员NF E2、c jun和c fos转录因子的诱导作用 ,探讨PNS在造血细胞内的信号传递途径 ,选用人粒系HL 6 0、红系K5 6 2、巨核系CHRF 2 88和Meg 0 14个细胞株作靶细胞 ,经PNS刺激处理后提取细胞核蛋白 ,分别用人抗NF E2、c jun和c fos抗体与核蛋白作Wes ternblot杂交试验 ,并用3 2 P标记的具有与NF E2、c jun和c fos转录因子共同结合位点的AP 1双链寡核苷酸探针作电泳带移动阻滞试验 (EMSA)。结果表明 :①Westernblot显示PNS诱导HL 6 0 ,K5 6 2 ,CHRF 2 88和Meg 0 14株细胞的NF E2和c jun转录因子蛋白水平分别是未经处理细胞的 1.5 - 2 .5倍和 2 .0 - 3.0倍。诱导的c fos蛋白在K5 6 2 ,CHRF 2 88和Meg 0 13株细胞分别提高 2 .0 - 3.0倍 ,但HL 6 0细胞的c fos在PNS处理后无明显变化 ;②EMSA显示AP 1转录调控蛋白与DNA结合复合物的特异性条带 ,经PNS诱导后 4株细胞的AP 1蛋白与DNA复合物条带密度明显高于未经处理的细胞。结论 :PNS在造血细胞内对基因的转录调控涉及到AP 1家族转录因子成员NF E2、c jun和c fos,通过诱导这些转录因子量的增加 ,与特异性DNA促进子结合的活性增高而调控与细胞增殖分化相关的基因的表达。
To observe the effects of Panax Notoginosides (PNS) on up-regulation of AP-1 family transcription factors NF-E2, c-jun and c-fos for exploring intracellular signal pathway of PNS in hematopoietic cells, four human hematopoietic cells lines including myeloid HL-60, erythroid K562, megakaryoid CHRF-288 and Meg-01 were incubated in the presence of PNS for 14 days. The nuclear protein of cells were extracted and analyzed by Western blot with antibodies against NF-E2, c-fos and c-jun. Electrophoretic mobility shift assay (EMSA) was performed by using 32 P labeled AP-1 consensus oligonucleotide which contains binding site for NF-E2, c-jun and c-fos. The results showed that the transcription factors NF-E2, c-jun and c-fos of AP-1 family could be induced by PNS. Western blot demonstrated that the nuclear protein of both NF-E2 and c-jun in four cell lines treated by PNS were increased by 1.5-2.5-and 2.0-3.0-fold over untreated cells respectively. The c-fos protein in three cell lines of K562, CHRF-288 and Meg-01 was also elevated by 2.0-3.0-fold respectively, while c-fos protein in HL-60 cells was no detectable difference after PNS treatment. EMSA results in four cell lines indicated that AP-1 binding activity initiated by PNS was apparently elevated to form higher density band of AP-1-DNA complex. In conclusion, the intracellular transcription regulation initiated by PNS was involved in transcription factors NF-E2, c-jun and c-fos of AP-1 family members, which could play an important role in the up-regulation of genes expression related to proliferation and differentiation of hematopoietic cells.
出处
《中国实验血液学杂志》
CAS
CSCD
2004年第1期16-19,共4页
Journal of Experimental Hematology
基金
国家自然科学基金资助项目
编号 3 0 0 70 93 3
