转化生长因子β1对系膜细胞内1型纤溶酶原激活物抑制物基因组蛋白乙酰化修饰的影响

Role of transforming growth factor 151 in the process of histone acetylate modification of plasminogen activator inhibitor 1 gene in mesangial cells

目的探讨转化生长因子β1（TGF-β1）对1型纤溶酶原激活物抑制物（PAI-1）基因启动子及转录区域内组蛋白尾部乙酰化修饰的影响。方法采用染色质共免疫沉淀与实时定量PCR技术，观察高糖及TGF-β1对PAI-1基因启动子及转录区组蛋白H3赖氨酸残基9位乙酰化（H3K9Ac）修饰的影响；并应用共免疫沉淀方法探讨转录因子CREB结合蛋白（CBP）与Smad3、Spl蛋白之间的相互作用。结果在PAI-1基因启动子的4个区域内，TGF-131（10μg／L）刺激显著增加了P1、P2、P3区域的H3K9Ac修饰（均P〈0．05），而在距离转录起始点较远的P4区域，H3K9Ac修饰水平没有明显改变；而在PAI-1基因的转录区，TGF-β1刺激显著增加了T1区域的H3K9Ac修饰（P〈0．05），而T2区没有明显改变。高糖刺激引起系膜细胞内PAl-1基因的表达水平及其启动子P1区域H3K9Ac修饰显著增加（P〈0．05），应用TGF-βl中和性抗体预处理显著抑制了高糖引起的PAI-1基因启动子区H3K9Ac修饰（P〈0．01）。TGF-β1刺激能够显著诱导Smad3、CBP蛋白结合在P1、P2、P3区域（均P〈0．05）；同时，TGF-β1刺激能够诱导CBP、Spl与Smad3蛋白的结合，进而引起H3K9Ac修饰。结论TGF-β1能够诱导PAI-1基因启动子与转录起始区发生H3K9Ac修饰，促进Smad3与Spl及CBP蛋白相互结合，促进PAI-1基因的转录表达。

Objective To explore the effect of transforming growth factorβ1 （TGF-β1） on epigenetie histone lysine aeetylation in the plasminogen activator inhibitor 1 （PAl- 1） promoter and transcribe regions in glomerular mesangial cells （GMCs）. Methods Chromatin immunopreeipitafion assay and real-time quantitative PCR were used to detect Histone3K9 acetlattion （H3K9Ae） in the PAI-1 promoter and transcribe regions induced by TGF-β1 and high glucose. Immunopreeipitation was also used to see the cooperation of Smad3, CBP and Spl proteins. Results In the four target regions of PAI-1 promoter, TGF-β1 treatment enhanced H3K9Ae at Pl, P2 and P3 in GMCs （P 〈 0.05）, but no change was seen in the P4 region which was far from the transcription starting site. TGF-β1 obviously induced H3K9Ac in the T1 transcribe region of PAI- l instead of T2 （P 〈 0.05）. High glucose increased PAI- 1 mRNA expression and H3K9Ac around P1 promoter region （P 〈 0.05）. TGF-β1 neutralizing antibody abrogated high glucose-induced H3K9Ac at PAI-1 promoter （P 〈 0.01）. TGF-[31 treatment could recruit Smad3 and CBP protein binding to the PAI- 1 promoter regions （P1, P2, P3）, and induce their cooperation in GMCs, which were responsing to TGF-β 1 associated H3K9Ac. Conclusion TGF-~ 1 can induce H3K9Ac in the promoter and transcribe regions of PAI- 1, promote Smad3 recruition and cooperation with Spl and CBP, which are associated with PAI-! gene＇ s regulation in GMCs.