背景：前期研究表明，姜黄素对细胞氧化应激炎症有重要作用。 目的：拟进一步阐明姜黄素在血管内皮细胞病理性炎症反应中的生物学作用及机制。 方法：以人血管内皮细胞为细胞模型，应用肿瘤坏死因子α(10μg/L)作为刺激因子，姜黄素(0，50，100µmol/L)孵育24 h作为干预治疗组。应用HRP标记的BSA检测内皮细胞通透性，应用罗丹明标记的鬼笔碱染色检测F-actin 变化，酶联免疫吸附实验检测细胞分泌白细胞介素1β的变化，进一步通过免疫荧光染色检测细胞内核转录因子κB蛋白表达和转位；应用Western blot方法检测炎症小体相关基因NRLP-3和caspase-1的表达。 结果与结论：HRP-BSA检测提示，姜黄素可呈剂量依赖性对抗刺激引起的血管内皮细胞通透性增加。同时，抑制F-actin应力纤维的形成。ELISA检测发现，姜黄素治疗后，可呈剂量依赖性抑制肿瘤坏死因子α刺激引起的血管内皮细胞白细胞介素1β分泌。同时，免疫荧光分析证实，肿瘤坏死因子α刺激后，对照组血管内皮细胞中核转录因子κB表达持续增高且发生明显的入核转位现象，而100μg/L姜黄素干预组细胞中炎症递质转录因子核κB表达定位无明显改变。Western blotting结果证实，炎症小体调控基因NRLP3和caspase-1表达在姜黄素干预组受到明显抑制。结果表明姜黄素可通过减少核转录因子κB 的表达对抗肿瘤坏死因子α刺激引起的炎症小体活化和白细胞介素1β分泌，对抗细胞病理性炎症损伤发生。%BACKGROUND:Previous studies have shown that, curcumin plays a crucial role on the inflammation in cells caused by oxidative stress. OBJECTIVE:To elucidate the biological effect and mechanism of curcumin in the pathological inflammation reaction in vascular endothelial cells. METHODHuman vascular endothelial cells were taken as the cellmodels. Tumor necrosis factor (10μg/L) treatment was used to induce the inflammation of cells. Curcumin (0, 50, 100μg/L) treatment for 24 hours was used to intervene the cells. The intercellular hyperpermeability of the vascular endothelial cellmonolayers was examined by HRP-conjugated bovine serum albumin, and intercellular filamentous actin stress fiber formation was examined by rhodamin-phal oidin staining. ELISA assay was used to detect the secretion of interleukin-1βin vascular endothelial cells. Immunoflurensece staining was applied to investigate the expression and translocalization of nuclear factor-κB. Western blot analysis reflected the expression of NRLP3 and caspase-1. RESULTS AND CONCLUSION:HRP-bovine serum albumin detection results showed that, curcumin inhibited the intercellular hyperpermeability of the vascular endothelial cellmonolayers and the formation of robust intercellular filamentous actin in a dose-dependent manner. ELISA assay showed that curcumin protected vascular endothelial cells against tumor necrotic factor-α-induced interleukin-1βsecretion in a dose-dependent manner. Meanwhile, the nuclear factor-κB expression was increased and the translocation of nuclear factor-κB into the nuclei was obviously seen in vascular endothelial cells induced by tumor necrosis factor-α, but the translocation was not changed in 100μg/L curcumin-treated cells. Furthermore, western blot analysis revealed that the expression of NRLP3 and caspase-1 were inhibited in curcumin-treated cells. Curcumin can inhibit tumor necrosis factor-α-induced activation of inflammasome and secretion of interleukin-1βin vascular endothelial cells by down-regulating the expression of nuclear factor-κB, thus prevent pathological inflammatory injury in cells.
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