中文摘要：

背景血管内皮细胞的增生和迁移是血管发生的主要环节。新近的研究发现，热休克蛋白B6（HspB6）可促进多种与血管新生相关因子的分泌，导致病理状态下新生血管的发生，研究HspB6中和性抗体对人脉络膜血管内皮细胞增生、迁移和管腔形成能力的影响对于脉络膜新生血管相关疾病的靶向治疗具有重要意义。目的探讨HspB6中和性抗体对人脉络膜血管内皮细胞管腔形成能力的影响及其机制。方法对人脉络膜血管内皮细胞株进行培养和传代，取对数生长期细胞用于实验。应用逆转录PCR（RT—PCR）和流式细胞术检测HspB6mRNA及其蛋白在人脉络膜血管内皮细胞中的表达。将基质胶铺于96孔板中，凝固后加入细胞悬液，细胞密度为2×10^4个／孔，将不同质量浓度（100μg／L、500μg／L）HspB6中和性抗体分别加入培养孔，未加入HspB6中和性抗体的细胞作为对照组（0μg／LHspB6中和性抗体组），每组设3个复孔。细胞孵育12h后，采用体外三维成型法检测各组完整管腔形成的数量，以评价HspB6中和性抗体对人脉络膜血管内皮细胞的管腔形成能力。96孔板培养人脉络膜血管内皮细胞，培养液中加入不同质量浓度（0、50、100、500μg/L）HspB6中和性抗体孵育24h，然后采用细胞计数试剂盒-8（CCK-8）检测细胞的增生值。采用Transwell小室法检测HspB6中和性抗体作用24h后各组穿过小室膜的“贴壁”细胞数目，以评估人脉络膜内皮细胞的增生和迁移情况。收集HspB6中和性抗体干预后的人脉络膜血管内皮细胞，流式细胞术检测HspB6中和性抗体作用后各组人脉络膜血管内皮细胞的凋亡率。结果人脉络膜血管内皮细胞在基因和蛋白水平均可表达HspB6分子。0、100、500μg／LHspB6中和性抗体组管腔形成数目分别为（67．25±5．75）、（60．39±6．41）和（39．76±10．73）个／视野，总体比较差异有?

英文摘要：

Background The proliferation and migration of vascular endothelial cells is a primary link during angiogenesis. Studies showed that heat shock protein B6 （HspB6） promotes the secretion of multiple angiogenesis-related factors and therefore leads to neovascularization. Understanding the effects of neutralizing HspB6 antibody on the biological behavior of human choroidal vascular endothelial cells has an important significance in the target treatment of choroidal neovacularization diseases. Objective This study was to address the role and mechanism of neutralizing HspB6 antibody in tube formation of human choroidal vascular endothelial cells. Methods Human choroidal vascular endothelial cell line was normally cultured and harvested for total RNA extraction. Expressions of HspB6 mRNA and protein in human choroidal vascular endothelial ceils were detected by reverse transcription PCR （RT-PCR） and flow cytometry （FCM）. The ceils were seeded on 96-well plate covered with matrigel at the density of 2~ 104/hole. Then the neutralizing HspB6 antibody at the concentration of 100μg/L and 500 μg/L was added into the medium respectively,and the control cells were set without the addition of HspB6 antibody. The number of capillary tubes was calculated 12 hours after culture by three-dimensional matrigel assay. In addition,0,50,100,500μg/L of neutralizing HspB6 antibody were added into the cell medium separately for 24 hours,cell counting kit-8 （CCK-8） method was employed to assay the inhibitory rate（IR） of the cells. Transwell test was used to count the cell number across chamber membrane for the evaluation of migration ability of the cells. The apoptosis of the cells was assayed by FCM. Results Both HspB6 mRNA and protein were expressed on human choroidal vascular endothelial cells. The number of capillary tube formation of human choroidal vascular endothelial cells was （ 67.25±5.75 ） , （ 60.39±6.41 ） and （ 39.76±10. 73 ） /field in the 0,100 and 500 μg/L neutralizing HspB6 antibody groups