| 摘要: |
| 目的:构建携带SAA3启动子的IκBα表达载体,观察其对NF-κB活性及脂多糖(LPS)诱导的小鼠炎症反应的影响,探讨脓毒症的治疗方法。方法:离体细胞实验：离体混合培养小鼠肝细胞和库普弗细胞,分为正常组、LPS处理组和LPS+基因转染组,LPS注射24 h后,测定各组细胞上清AST、ALT、LDH和TNF-α、IL-6水平。小鼠在体实验：(1)小鼠随机分为正常组、LPS处理组和LPS+基因转染组3组(n=10),小鼠腹腔注射250 μg LPS或等量生理盐水,24 h后处死,取血清和肝组织测定TNF-α、IL-6水平。(2)小鼠随机分为LPS处理组和LPS+基因转染组(n=21),0、48 h二次注射150 μg LPS,首次注射后不同时点(0、2、24、48、50、72、96 h)取肝组织测NF-κB和IκBα活性,以0 h测得值作为正常对照。(3)小鼠随机分为LPS处理组和LPS+基因转染组(n=20),腹腔注射350 μg LPS后,继续饲养96 h,观察各时点(0、12、24、36、48、72、96 h)小鼠生存率。结果:与LPS组相比,LPS+基因转染组共培养细胞上清中AST、LDH和TNF-α、IL-6水平均降低,但仍高于正常组(P<0.05)。与LPS组相比,LPS+基因转染组肝组织、血清中TNF-α、IL-6水平降低(P<0.05)；与LPS组相比,LPS+基因转染组2、24、50、72 h时NF-κB活性明显降低,但高于正常对照(P<0.05)；LPS刺激72、96 h时LPS+基因转染组小鼠的存活率高于LPS组(P<0.05)。结论:以SAA3为启动子,IκBα靶基因能够在肝脏表达,并可有效动态抑制内毒素诱导的小鼠肝脏或全身炎症过激反应。 |
| 关键词: NF-κB；IκBα；基因疗法 炎症 脂多糖类 |
| DOI：10.3724/SP.J.1008.2009.01240 |
| 投稿时间：2008-10-07修订日期：2008-11-13 |
| 基金项目:上海市启明星人才培养基金 |
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| Inhibitory effect of IκBα gene on LPS-induced inflammation in mice |
| LI Quan,YU Wei-feng*,QIU Bi-jun,ZHANG Jin-min,FEI Guo-xiong |
| (Department of Anesthesiology,Eastern Hepatobiliary Hospital,Second Military Medical University,Shanghai 200438,China) |
| Abstract: |
| Objective:To construct IκBα vector carrying SAA3 promoter and to observe its effect on NF-κB activity and LPS-induced inflammation,so as to lay a foundation for treatment of sepsis.Methods: Mouse liver cells and kupffer cells were co-cultured and were divided into three groups:control group,LPS group and LPS+ gene transfer group.Twenty-four hours after LPS injection,the levels of AST,ALT,LDH,IL-6 and TNF-α were measured in the supernatants of the each group.For animal experiments: (1) Mice were divided into three groups:control group,LPS group and LPS+gene transfer group(n=10).The levels of TNF-α and IL-6 were measured in the serum and liver tissues 24 hours after intraperitoneal injection of 250ug LPS or saline.(2) Mice were also divided into two groups:LPS group and LPS+gene transfer group(n=21).Mice were injected with 150 μg LPS twice at 0 and 48 h,then the activities of NF-κB and IκBα in the liver were measured at 0,2,24,48,50,72,and 96 h after the first injection.The values at 0 h were taken as control group.(3) Mice were also divided into another two groups:LPS group and LPS+gene transfer group(n=20).The survival rates of animals were observed at 0,2,24,48,50,72,and 96 h after injection of 350 μg LPS.Results: Compared with LPS group,the levels of AST,LDH,TNF-α and IL-6 in the culture supernatants of LPS+gene transfer group were decreased,but were still higher than those in the control group(P<0.05).Compared with LPS group,the levels of TNF-α and IL-6 in the liver tissues and sera of LPS+gene transfer group were significantly decreased(P<0.05).Compared with LPS group,the activity of NF-κB in the liver tissues of LPS+gene transfer group were decreased,but was still significantly higher than that of the control group(P<0.05).Compared with LPS group,LPS+gene transfer group had higher survival rate at 72 and 96 h(P<0.05).Conclusion: IκBα gene can be expressed in the liver with SAA3 promoter,and transfection of IκBα can effectively inhibit endotoxin-induced liver and general inflammation. |
| Key words: NF-κB IκBα gene therapy inflammation lipopolysaccharides |
