西红花酸对晚期糖化终产物诱导内皮细胞晚期糖基化终产物受体表达的抑制作用

目的: 研究西红花酸对晚期糖基化终产物(AGE)诱导牛内皮细胞(BEC)中晚期糖基化终产物受体(RAGE)mRNA表达的抑制作用,并探讨其可能机制。方法: 不同浓度的西红花酸(1、0.1、0.01 μmol/L)预孵BEC细胞12h后,用AGE(100 mg/L)刺激细胞12h,RT-PCR法测定RAGEmRNA的表达水平;ELISA法测定细胞间黏附分子-1(ICAM-1)的表达;试剂盒分别检测胞外超氧阴离子和硫代巴比妥酸反应产物(TBARS)浓度;同时,还用2,7-二氯荧光素(DCFH)测定了胞内H2O2的浓度,并用罗丹明123(Rh123)荧光法及MTT法分别检测细胞线粒体膜电位(MMP)水平和其琥珀酸脱氢酶(MSDH)的活性。结果: 与AGE模型组相比,西红花酸能显著抑制RAGEmRNA的表达(P<0.05),降低胞外超氧阴离子和TBARS(P<0.01或P<0.05)及胞内H2O2水平;结果还显示,西红花酸能提高细胞MMP水平和MSDH活性。对ICAM-1蛋白表达也有抑制作用,且呈时间和剂量依赖性。结论: 西红花酸可能通过清除AGE与RAGE结合产生的活性氧(ROS)来抑制RAGEmRNA的高表达。提示西红花酸对糖尿病血管病变有潜在的治疗价值。

Crocetin reduces expression of receptor for advanced glycation end products (RAGE) on endothelial cells induced by AGE

AIM: To investigate the effects of crocetin on receptor for advanced glycation end products (RAGE) mRNA expression in bovine endothelial cells (BEC) induced by advanced glycation end products (AGE) and the possible mechanisms involved.METHODS: The BEC were preincubated with crocetin(1, 0.1 and 0.01 μmol/L) for 12 h, then exposed to AGE (100 mg/L).The RAGE mRNA expression was detected by RT-PCR analysis, the intercellular adhesion molecule-1 (ICAM-1) was measured by ELISA. The extracellular superoxide ion and thiobarbituric acid reactive substances (TBARS) were assessed with superoxide ion kit and colorimetric assay, respectively. The intracellular H₂O₂ was also detected using the probe 2, 7-dichlorofluorescein (DCFH).The mitochondrial membrane potential(MMP) and mitochodrial Succinate dehydrogenase(MSDH) were analyzed by the retention of rhodamine123 (Rh123) and MTT.RESULTS: Compared with AGE Group, crocetin was able to significantly reduce RAGE mRNA expression (P<0.05), decrease the content of super anion and TBARS in supernatant media (P<0.01 or P<0.05) and H₂O₂ in cells (P<0.05).Spontaneously, the MMP and the activity of MSDH were improved.Furthermore, the expression of ICAM-1 protein was suppressed by crocetin in a time-and dose-dependant manner.CONCLUSION: These results demonstrated that crocetin could inhibit RAGE over-expression in AGEexposed BEC by suppressing the generation of reactive oxygen species active oxygen(ROS).This study suggested that crocetin might have beneficial effects in the treatment of diabetic vascular complications.