组蛋白去乙酰化酶抑制剂保护烧伤后血管内皮细胞屏障的作用和机制研究

The protection of vascular endothelial barrier (VEB ) from damage caused by major burn injury is of great importance in the prevention and treatment of shock and tissue edema. Emerging evidence suggests that hypoxia and hyperthemia activate the hypoxia-inducible factor-1(HIF-1)/heat shock protein 90 (HSP90) signaling pathway which promotes the transcription of downstream gene vascular-endothelial-barrier regulating-factors which have been demonstrated to be strong regulators involved in endothelial skeleton loose and systolic dysfuction. Histone deacetylase inhibitors (HDACIs), have recently been shown to be able to prevent the skeleton protein contraction and inhibit HIF-1/HSP90, but their protective effects and mechanism of burn-induced VEB damage remain unknown. We assume that the HDACIs may possibly protect VEB function through inhibiting HIF-1 alpha/HSP90 function leading to stabilizing endothelial skeleton. The objectives of this research are: 1. to confirm whether or not HDACIs can protect endothelial barrier function from hypoxia/hyperthemia/burn serum-induced injury in a cultivated endothelium; 2. to investigate the mechanism by down-regulatoery effects on and the acetylation levels of HIF-1 and its stimulus molecule, as well as the synergy of HSP90 as molecular chaperone; and 3. to explore the therapeutic potential of HDACIs in treating severe burn-induced plasmexhidrosis and tissue edema due to VEB dysfunction.

保护血管内皮屏障（VEB）功能，对于减轻烧伤引起的血浆渗出和组织水肿、防治休克有重要意义。烧伤组织缺氧和高温能激活缺氧诱导因子1（HIF-1）/热休克蛋白90(HSP90)信号通路，增强下游靶基因血管内皮屏障调节因子的转录，增加肌球蛋白轻链磷酸化，导致内皮骨架蛋白收缩，损害VEB功能。以往研究表明，组蛋白去乙酰化酶抑制剂（HDACI）能抑制HIF-1/HSP90，但其对烧伤VEB损害的作用及机制尚不清楚。我们假设HDACIs能通过下调HIF-1α/HSP90，稳定内皮细胞骨架，保护VEB功能。研究目标：①采用内皮细胞低氧、高温培养和烧伤血清刺激，确认HDACIs对VEB有保护作用; ②研究HDACI通过下调HIF-1α及其共刺激分子的乙酰化水平和抑制HSP90的分子伴侣作用，降低HIF-1的功能、保护VEB的机制。③制作严重烧伤动物模型验证HDACIs减少渗出和补液量、防治休克的效果。

项目研究背景：保护血管内皮屏障功能，对于减轻烧伤引起的血浆渗出和组织水肿、防治休克有重要意义。烧伤组织缺氧和高温能激活缺氧诱导因子1（HIF-1）和热休克蛋白90(HSP90)信号通路，增强下游靶基因血管内皮屏障调节因子的转录，增加肌球蛋白轻链磷酸化，导致内皮骨架蛋白收缩，损害内皮屏障功能。以往研究表明，组蛋白去乙酰化酶抑制剂（HDACIs）能抑制HIF-1和HSP90，但其对烧伤血管内皮屏障损害的作用及机制尚不清楚。主要研究内容：①采用烧伤动物模型和缺氧/烧伤血清刺激内皮细胞模型，确认HDACIs对血管内皮屏障具有保护作用;②研究HDACIs通过下调HIF-1α的乙酰化水平和抑制HSP90的分子伴侣作用，降低HIF-1功能、保护血管内皮屏障的机制；③制作严重烧伤休克大动物模型验证HDACIs减少渗出和补液量、防治休克的效果。重要研究结果：①HDACIs(丙戊酸钠或/和丁酸钠)不仅能有效抑制严重烫伤大鼠模型肺微血管内皮通透性增加，减轻脏器组织水肿和炎症损害；而且能显著减轻缺氧或/烫伤血清刺激引起的肺微血管内皮细胞通透性增加和屏障功能损伤。②丙戊酸钠或/和丁酸钠能恢复血管内皮细胞组蛋白乙酰化水平，抑制HIF-1及其分子伴侣HSP90、减少HIF-1α的下游靶基因VEGF和MLCK的表达，增加血管内皮细胞连接骨架蛋白的稳定性。③采用50%TBSAIII度烧伤大鼠和比格犬模型进一步证明HDACIs（NK-HDAC-1、丁酸钠或血必净）能通过抑制HIF-1信号通路、保护血管内皮屏障，减少烧伤休克血浆渗出和补液量、显著改善血流动力学指标和脏器功能。主要科学意义：本项目研究为HDACIs用于防治创（烧）伤、缺血、炎症引起的血管内皮屏障损害提供了新的理论依据、研究方法和潜在药物，对平时和战时创、烧伤休克救治均有重要意义。