咪唑克生对内毒素攻击小鼠体内及体外活化巨噬细胞炎性介质释放的抗炎效应  

The anti-inflammatory effects of idazoxan on inflammatory mediator release in endotoxin-challenged mice in vivo and activated macrophages in vitro

作　　者：李香琴[1] 朱俊宇[2] 马娓 罗莉[2] 梁华平[1] 

机构地区：[1]成都医学院基础医学院,四川成都610500 [2]第三军医大学大坪医院野战外科研究所一室创伤、烧伤与复合伤国家重点实验室,重庆400042

出　　处：《中华危重病急救医学》2016年第5期445-449,共5页Chinese Critical Care Medicine

基　　金：国家重点基础研究发展计划（973）项目（2012CB518102）;国家重点实验室项目（BWS11J038）

摘　　要：目的观察咪唑克生（IDA）对内毒素脂多糖（LPS）攻击小鼠体内以及体外活化巨噬细胞的抗炎效应，探讨其可能的分子机制。方法体内实验：将30只成年雄性C57BL／6小鼠按随机数字表法分为对照组、模型组及IDA低、中、高剂量组（IDA—L、IDA—M、IDA—H组），每组6只。采用腹腔注射LPS10mg／kg制备炎症模型，对照组注射等量生理盐水；IDA各组制模同时相应注射IDA0．3、1．0、3．0mg／kg进行干预。于制模后6h取血备检。体外实验：收集20只成年雄性C57BL／6小鼠原代腹腔巨噬细胞，分为空白组、LPS组（10mg／L）及LPS＋IDA—L、IDA—M、IDA—H组（10mg／LLPS＋5、25、100μmol／LIDA），于24h收集培养上清备检。检测方法：采用酶联免疫吸附试验（ELISA）测定血清肿瘤坏死因子-α（TNF—α）、白细胞介素-6（IL-6）、单核细胞趋化因子-1（MCP-1）及一氧化氮（NO）含量；采用蛋白质免疫印迹试验（Western Blot）测定IDA对巨噬细胞核转录因子-κB（NF-κB）表达的影响。结果①体内实验：模型组血清TNF-α（ng／L：403．96±40．98比17．50±8．68）、IL-6（ng/L：61400．31±7826．61比2436．30±448．89）均较对照组显著升高（均P〈0．01）；IDA能剂量依赖性地抑制炎性因子的升高，以IDA—H组尤甚[TNF—α（ng／L）：170．09±28．53比403．96±40．98，IL-6（ng／L）：16570．81±1083．65比61400．31±7826．61，均P〈0．01]。②体外实验：与空白组比较，LPS组巨噬细胞分泌炎性因子量明显增加（TNF—α（ng／L）：7259．14±320．70比28．50±27．08，IL-6（ng／L）：14809．60±5852．73比1113．47±465．53，MCP-1（ng，L）：20847．37±1788．33比447．37±395．69，NO（μmol／L）：1900．00±144．31比603．03±102．18，均P〈0．01]；IDA能剂量依赖性地抑制上述因子的分泌，以LPS＋IDA—H组作用尤甚[TNF—α（ng／L）：784．40±281．90比7259．14±320Objective To study the anti-inflammatory effects of idazoxan （IDA） on endotoxin lipopolysaccharide （LPS） challenged mice in vivo and activated macrophages in vitro, and explore its potential molecular mechanisms. Methods To do the experiments in vivo, 30 adult male C57BL/6 mice were randomly divided into control group, model group, and low, medium and high doses IDA groups （IDA-L, IDA-M, and IDA-H groups）, n = 6 in each group. The inflammatory model was reproduced by intraperitoneal injection of LPS 10 mg/kg, and the control group was injected with the same amount of normal saline. The IDA groups received LPS （10 mg/kg） and IDA 0.3, 1.0 and 3.0 mg/kg, respectively. The blood samples of mice in each group were collected at 6 hours after the reproduction of the model.For the in vitro experiments, primary peritoneal macrophages were collected from 20 adult male C57BL/6 mouse cells and they were divided into control group, LPS group （10 mg/L） and LPS＋IDA-L, IDA-M, IDA-H groups （10 mg/L LPS ＋ 5, 25, 100 μmol/L IDA, respectively）. Cell culture supernatants were collected at 24 hours after the reproduction of the model. Detection methods： enzyme linked immunosorbent assay （ELISA） was used to determine the levels of serum tumor necrosis factor-or （TNF-α）, interleukin-6 （IL-6）, monocyte chemotactic protein-1 （MCP-1） and nitric oxide （NO）. Western Blot was used to determine the effect of IDA on the expression levels of nuclear factor-κB （NF-κB） in macrophages. Results ① For the in vivo experiment, the serum levels of TNF-α and IL-6 were significantly elevated in the model group as compared with those in the control group [TNF-α （ng/L）： 403.96 ± 40.98 vs. 17.50 ± 8.68; IL-6 （ng/L）： 61400.31 ± 7826.61 vs. 2 436.30 ±445.89; both P 〈 0.01]. IDA treatment could inhibit the elevation of inflammatory cytokines in a dose-dependent manner, with the most significant decrease in LPS＋IDA-H group [TNF-α （ng/L）： 170.09±28.53 vs. 403.96±40.98, IL-6

关 键 词：咪唑克生 内毒素 巨噬细胞 抗炎作用 分子机制 

分 类 号：R965[医药卫生—药理学]