尿毒症毒素硫酸盐对甲酚对血管内皮细胞的体外作用

目的探讨尿毒症毒素硫酸盐对甲酚(PCS)对人脐静脉内皮细胞(HUVECs)的毒性作用及机制。方法体外培养HUVECs,不同浓度对甲酚硫酸钠盐作用HUVECs,WST-1法检测细胞增殖;Matrigel法检测细胞成血管能力;特异性荧光探针2’,7’-二氯荧光素二乙酸酯(DCFH-DA)检测细胞活性氧自由基(ROS)水平;实时定量聚合酶链反应(Real-time PCR)及免疫印迹法(Western blotting)检测细胞中炎症因子及黏附分子表达情况。结果硫酸盐对甲酚对HUVECs增殖及成血管能力未有显著影响,分子机制研究发现,其能显著诱导细胞中ROS水平上调并促进其分泌炎症因子白细胞介素(IL)-1β、肿瘤坏死因子-a(TNF-a)及细胞间黏附分子-1(ICAM-1)。结论尿毒症毒素硫酸盐对甲酚可以加剧血管内皮细胞氧化应激及炎症反应状态,提示其可能参与慢性肾病患者动脉粥样硬化的发生发展。

Effects of the uremic toxin p-cresylsulfate on human umbilical vein endotheilial cell function in vitro

Objective To investigate the effect of a uremic toxin, p-cresylsulfate, on human umbilical vein endothelial cells (HUVECs) function. Methods HUVECs were incubated with various concentrations of p-cresylsulfate (10, 40, 80, and 160 mg/L) for up to 72 hours. Cell proliferation was determined using WST-1 assay. Tube formation assay was used to evaluate HUVECs function in vitro. Reactive oxygen species (ROS) production in HUVECs was measured by fluorescence microscopy. Inflammatory factor and adhesion molecule expressions were assessed by real time quantitative PCR(Real-time PCR) and Western blotting analysis. Results P-cresylsulfate had no anti-proliferative effects on HUVECs. Cells treated with concentrations of p-cresylsulfate in the range found in CKD patients had no observable impairment on tube formation. In contrast, p-cresylsulfate partially increased ROS production in HUVECs, while quantitative Real-time PCR and Western blotting revealed p-cresylsulfate treatment increase interleukin(IL)-1β, tumor necrosis factor(TNF-a), and intercelluar adhesion molecule\|1(ICAM-1) expressions in HUVECs. Conclusion Our study revealed that a uremic toxin, p-cresylsulfate, have pro-oxidant and pro-inflammatory effects on HUVECs, which indicates that p-cresylsulfate may be among the factors involved in the high incidence of atherosclerosis in patients with chronic kidney disease.