| 刘珍珍,肖 轶,毛萧萧,段 琼,杨天伦.BET蛋白通过调控NF-κB通路参与炎症基因转录[J].,2017,17(23):4456-4461 |
| BET蛋白通过调控NF-κB通路参与炎症基因转录 |
| BET Bromodomain Involves in Inflammatory Genes Transcription via Regulation of NF-κB Signaling Pathway |
| 投稿时间：2017-03-03 修订日期：2017-03-29 |
| DOI：10.13241/j.cnki.pmb.2017.23.012 |
| 中文关键词: JQ1 BET蛋白 NF-κB 炎症 |
| 英文关键词: JQ1 BET NF-κB Inflammation |
| 基金项目:国家自然科学基金项目(81570334)；中南大学中央高校基本科研业务费专项资金项目（2015zzts298） |
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| 中文摘要: |
| 摘要 目的：探讨特异性抑制溴结构域(bromodomain and extra-terminal, BET)蛋白对血管内皮细胞激活与早期动脉粥样硬化形成的作用及其分子机制。方法：1.原代分离培养脐静脉内皮细胞和小鼠心脏血管内皮细胞后用肿瘤坏死因α(TNFα）刺激模拟炎症过程，以小分子化合物JQ1特异性抑制BET蛋白，分组如下：(1)对照组；(2)TNFα(25 ng/mL）处理组；(3) TNFα+JQ1处理组。采用Realtime-PCR及流式细胞术检测各组细胞炎症因子mRNA及蛋白水平的表达，采用5XκB荧光素酶报告基因检测各组核转录因子kappa B(NF-κB）转录活性。2.LDL受体基因敲除(LDLR-/-）小鼠随机分为2组：JQ1组(n=8，JQ1腹腔注射，50 mg/kg，每天一次）和对照组(n=8，DMSO溶媒组），同时给予高胆固醇饮食8周，采用免疫组化方法检测主动脉弓部血管细胞黏附分子-1(VCAM-1）表达水平。结果：与对照组相比，TNFα组炎症因子mRNA、蛋白表达明显上调(P<0.01），使用JQ1干预后，炎症因子E选择素(E-selectin）、P选择素(P-selectin）、VCAM-1及白细胞介素-8(IL-8）mRNA及蛋白表达均明显下调(P<0.01）。LDLR-/-小鼠高脂饮食诱导8周后JQ1显著下调了主动脉弓部VCAM-1蛋白表达。5XκB荧光素酶报告基因结果显示，与TNFα(-)相比，TNFα(+)组荧光素酶报告基因活性增强，JQ1可以显著下调报告基因活性(P<0.01）。结论：BET蛋白通过调控NF-κB信号通路参与了血管内皮炎症基因转录；抑制BET蛋白下调了NF-κB目的基因表达从而减轻了内皮激活及高脂诱导的早期动脉粥样硬化病理改变。 |
| 英文摘要: |
| ABSTRACT Objective: To explore bromodomain and extra-terminal (BET) inhibition in the regulation of vascular endothelial cells activation and early atherosclerosis formation and its potential molecular mechanisms. Methods: 1. Human umbilical vein endothelial cells (HUVEC) and mouse heart endothelial cells (MHEC) were isolated, and tumor necrosis factor α (TNFα) was used to activate in- flammatory genes transcription in the presence or absence of JQ1, a specific BET inhibitor. The groups are as follows: (1)Normal control group; (2) TNFα(25 ng/mL）group; (3) TNFα+JQ1 group. The gene mRNA and protein expression of inflammatory cytokines were mea- sured by both real-time PCR and flow cytometry (FCM). 2. LDL receptor-deficient (LDLR-/-) mice were randomly divided into 2 groups: JQ1 group (n=8, JQ1intraperitoneal, 50 mg/kg, daily) and control group (n=8, DMSO, daily). After 8 weeks feeding with high cholesterol diet, vascular cell adhesion molecule-1 (VCAM-1) expression in aortic arch was measured by immunohistochemistry. The activity of nu- clear factor kappa B (NF-κB) signaling was monitored by 5XκB luciferase reporter assay in HEK293. Results: TNFα dramatically in- duced the mRNA and protein expression of inflammatory genes and JQ1 significantly downregulated the induction of them (E-selectin, P-selectin, VCAM-1, IL-8)(P<0.01). Immunohistochemistry detection indicated that JQ1 significantly downregulated the expression of VCAM-1 in aortic arch induced by 8 weeks high cholesterol diet feeding comparing to control group. In addition, BET bromodomain in- hibition downregulated TNFα upregulated NF-κB transcriptional activity (P<0.01). Conclusion: Our study demonstrated that BET bro- modomain was involved in NF-κB mediated inflammatory genes expression; inhibition of BET bromodomain suppressed vascular en- dothelial activation in vitro, and attenuated early atherogenesis in vivo. |
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