组蛋白甲基转移酶EZH2介导的Sp4活化在腹膜血管新生中的作用研究

VEGF is closely related with peritoneal angiogenesis in peritoneal ultrafiltration failure. Our previous study confirmed for the first timethat IL-6/sIL-6R-induced VEGF could be targeted by non-classical transcription factor Sp4 in human peritoneal mesothelial cells (HPMCs), but the mechanism was not clear. The latest reports that IL-6/sIL-6R-VEGF signaling pathway is following with EZH2 regulatory abnormalities in tumorigenesis. Our group also found that IL-6/sIL-6R can promote the high expression of EZH2 and blocking EZH2 can inhibit the expression of Sp4 in HPMCs, and tubular formation in human umbilical vein endothelial cells. It is speculated that Sp4 activation may be related to epigenetic regulation of EZH2. Thus, we will establish peritoneal ultrafiltration failure rat mode to observe EZH2 regulation of histone H3K27 methylation and DNA methylation changes and Sp4 activation in peritoneal neovascularization via ChIP, mDIP and other techniques. Then gene Transfection and interfering RNA targeting are used to block EZH2 to investigate the regulation mechanism of EZH2 in Sp4-VEGF induced by IL-6 / sIL-6R. These results may provide an effective therapeutic target for inhibiting peritoneal angiogenesis and delaying the progress of peritoneal dialysis.

VEGF与腹透超滤衰竭中腹膜血管新生密切相关。本课题组前期研究首次在人腹膜间皮细胞(HPMCs)中证实，IL-6/sIL-6R诱导的VEGF可被非经典转录因子Sp4所靶向，但机制未明确。最新报道，在肿瘤发生中IL-6/sIL-6R-VEGF信号通路伴EZH2调控异常。本课题组也发现IL-6/sIL-6R可促使EZH2高表达，阻断EZH2可抑制HPMCs中Sp4表达、人脐静脉内皮细胞的小管形成能力减弱。故推测，Sp4活化可能与EZH2的表观遗传调控相关。为此，本课题建立腹膜超滤衰竭大鼠模型，运用ChIP、mDIP等技术，观察腹膜血管新生中EZH2调节的组蛋白H3K27甲基化和DNA甲基化的变化及Sp4活化水平；进而应用基因转染、干扰RNA打靶等技术，阻断EZH2，探讨EZH2在IL-6/sIL-6R诱导Sp4-VEGF的调控机制，为抑制腹膜血管新生、延缓腹膜透析超滤衰竭进程提供有效治疗靶点。

腹膜血管新生是腹膜进展至超滤衰竭重要的病理生理过程。VEGF作为腹膜血管新生的关键因子，核转录因子Sp4调控VEGF生成的机制尚不完全清晰。本课题组前期研究首次在人腹膜间皮细胞(HPMCs)中证实，IL-6/sIL-6R诱导的VEGF可被非经典转录因子Sp4所靶向，但机制未明确。最新报道，在肿瘤发生中IL-6/sIL-6R-VEGF信号通路伴EZH2调控异常。本课题组也发现IL-6/sIL-6R可促使EZH2高表达，阻断EZH2可抑制HPMCs中Sp4表达、人脐静脉内皮细胞的小管形成能力减弱。但是，EZH2在腹膜血管新生Sp4-VEGF通路活化中的作用及其机制如何。本课题通过体内外研究阐明EZH2在Sp4介导的腹膜透析血管新生中的作用机制。我们的研究证实（1）EZH2参与间皮细胞中IL-6/sIL-6R诱导的Dnmt3b和Sp4诱导和血管生成；（2）EZH2通过Sp4启动子区域中CpG岛的Dnmt3b甲基化激活Sp4；（3）EZH2参与HUVECs中IL-6/sIL-6R诱导的VEGF产生和血管生成；（4）EZH2参与5/6肾切除腹膜透析大鼠腹膜血管的生成；（5）沙利度胺通过腹膜STAT3/SP4信号通路抑制血管生成。不仅进一步明确Sp4在IL-6/sIL-6R-VEGF通路中的重要作用及其机制，也为延缓和阻断腹膜血管新生提供全新理论基础，为研发特异性抗腹膜血管新生药物提供新的实验依据和靶标，最终为腹膜透析患者超滤衰竭的防治提供新途径。发表相关论文2篇，授权专利4项。