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姜黄素调控炎症对内皮细胞增殖凋亡的影响
引用本文:薛君力,代喆,邓浩华,陈小奇,孙家忠,钟绍,徐焱成. 姜黄素调控炎症对内皮细胞增殖凋亡的影响[J]. 中华糖尿病杂志, 2013, 0(9): 536-540
作者姓名:薛君力  代喆  邓浩华  陈小奇  孙家忠  钟绍  徐焱成
作者单位:[1]武汉大学中南医院内分泌科,430071 [2]江苏大学附属昆山医院,430071
摘    要:目的明确姜黄素对炎症诱导的血管内皮细胞损伤的保护机制。方法将人单核/巨噬细胞系(THP-1)分为空白对照组、高脂高糖组、高脂高糖+姜黄素预处理组(高脂高糖浓度为25mmol/L葡萄糖+500μmol/L棕榈酸),分别给予相应处理24h,更换培养基继续培养24h,利用酶联免疫吸附法(ELISA)检测上清及实时定量聚合酶链反应(RT-PCR)分析THP-1细胞内肿瘤坏死因子-α(TNF-α)、白介素-6(IL-6)的表达,同时将上清作用脐静脉血管内皮细胞(HUVECs),HUVECs分为空白对照组、空对照条件培养基组、高脂高糖条件培养基组、姜黄素处理条件培养基组,并行噻唑蓝法(MTT)检测HUVECs增殖、流式细胞仪检测凋亡,Westernblotting分析磷酸化细胞外调节蛋白激酶(p-ERK)及B淋巴细胞瘤-2(Bcl-2)蛋白表达。组间比较采用独立样本t检验,多组间比较采用单因素方差分析。结果高脂高糖组THP-1细胞内受体相互作用蛋白140(RIPl40)、TNF-α和IL-6mRNA表达及上清中炎症因子TNF-α和IL-6浓度明显高于空白对照组(t=8.55、9.44、9.73、16.01、19.22,均P〈0.05)。相对于高脂高糖组,姜黄素预处理组THP-1细胞内RIPl40、TNF-α和IL-6mRNA表达及上清中TNF-α和IL-6浓度明显下降(t=3.59、5.96、5.59、6.95、23.91,均P〈0.05);高脂高糖条件培养基组HUVECs细胞活力明显低于空对照条件培养基组(24h,1.22±0.07比1.85±0.14,t=6.58,P〈0.05;48h,1.72±0.02比2.49±0.09,t=10.08,P〈0.05),而姜黄素处理条件培养基组能够改善高脂高糖条件培养基对HUVECs增殖的抑制作用(24h,1.22±0.07比1.72±0.11,t=2.13,P〈0.05;48h,1.72±0.02比2.33±0.11,t=6.92,P〈0.05);与空对照条件培养基组比较,高脂高糖条件培养基组能够明显增加HUVECs凋亡率、p-ERK表达及降低Bcl-2表达(t=9.82、9.69、4.61,均P〈0.05),姜黄素处理条件培养基组与高脂高糖条件培养基组比较,下调RIPl40表达后能明显降低HUVECs凋亡率、p-ERK表达及增加Bcl-2表达(t=6.35、7.17、3.26,均P〈0.05)。结论姜黄素能够通过抑制高脂高糖诱导的炎症来调控HUVECs的增殖凋亡。

关 键 词:姜黄素  受体相互作用蛋白140  炎症  血管内皮细胞

Curcumin suppresses the effect of inflammation on proliferation and apoptosis of vascular endothelial cells
Affiliation:XUE Jun-li , DAI Zhe, DENG Hao-hua, CHEN Xiao-qi, SUN Jia-zhong, ZHONG Shao, XU Yan- cheng. (Department of Endocrinology, Zhongnan Hospital of Wuhan University, Wuhan 430071, China)
Abstract:Objective To investigate the protective role of curcumin in the dysfunctional vascular endothelial cells induced by inflammation. Methods THP-1 cells were pretreated with 20 μmoL/L curcumin under the condition of 25 mmol/L glucose and 500 μmol/L palmitic acid, while other groups received 25 mmol/L glucose and 500μmoL/L palmitic acid and dimethyl sulfoxide ( DMSO ) , seperately. After 24 h treatment, the real time Polymerase Chain Reaction (PCR) was explored to detect the mRNA expression of receptor interaction protein 140(RIP140) , tumor necrosis factor-α (TNF-α) and interleukin- 6 ( IL-6), while the concentration of TNF-α and IL-6 in the supernatant was detected by the enzyme-linked immunosorbent assay (ELISA). Human umbilical vein endothelial cells (HUVECs) were incubated with the supernatant from the THP-1 in each group. The methodology of 3-(4, 5-Dimethylthiazol-2-yl)-2, 5- diphenyhetrazolium bromide (MTT) assay, flow cytometry and western-blot was applied to investigate the proliferation, apoptosis and the protein expression of phosphorylated extracellular regulated protein kinases (ERK) and B-cell lymphoma 2 (Bcl-2). Results The mRNA expression of RIP140, TNF-α and IL-6 in the THP-1 cells and the concentration of TNF-α and IL-6 in the supernatant were significantly higher when treated with 25 mmol/L glucose and 500μmol/L palmitic acid, than those in the control group (t = 8.55, 9. 44, 9.73, 16.01, 19.22 with all P 〈 0. 05). In the groups treated with 25 mmol/L glucose and 500 μmol/L palmitic acid, the pretreatment with curcumin significantly decreased the mRNA expression of RIP140, TNF-α and IL-6 in the THP-1 cells and the concentration of TNF-α and IL-6 in the supernatant (t = 3.59, 5.96, 5.59, 6. 95, 23.91 with all P 〈 O. 05). At 24 h and 48 h treatment, compared with the control group, the supertant from the groups treated with 25 mmol/L glucose and 500 Ixmol/L palmitic acid significantly suppressed the proliferation of HUVECs (24 h : 1.22 ± 0. 07 vs 1.85 ± 0. 14, t = 6. 58, P 〈 0. 05 ; 48 h : 1.72 ± 0. 02 vs 2. 49 ± 0. 09, t = 10. 08, P 〈 0. 05 ), while the proliferation of HUVECs could be improved by the pretreatment with 20 μmol/L curcumin (24 h : 1.22 ± 0. 07 vs 1.72 ± 0. 11, t = 2. 13, P 〈 0.05 ; 48 h: 1.72 ± 0.02 vs 2. 33 ± 0.11, t = 6.92, P 〈 0.05). Compared with the control group, the incubation with the supertant from the groups treated with 25 mmol/L glucose and 500 /xmol/L palmitic acid significantly increased the apoptosis rate and the level of phosphated ERK, while decreased the Bcl-2 protein expression (t =9.82, 9.69,4. 61, all P 〈0. 05). Such effect could be reserved by the pretreatment with 20μmol/L curcumin ( t = 6. 35, 7. 17, 3.26, all P 〈 0. 05 ). Conclusion Curcumin could protect the vascular endothelial cells from the dysfunction induced by inflammation.
Keywords:Curcumin  Receptor interaction protein 140  Inflammation  Vascular endothelial cells
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