不同剂量胸腺肽-α1在肠毒素B与脂多糖复合诱导小鼠脓毒症模型中的免疫干预作用研究  

Research on the immune intervention of different dosages of thymosin α1 on septic mouse model induced by enterotoxin B and lipopolysaccharide

作　　者：吕宝谱 田磊[1] 董艳玲 孙艺青 孟庆冰[1] 张睿[1] 刘亮[1] 肖浩[1] 田英平[1] Lyu Baopu;Tian Lei;Dong Yanling;Sun Yiqing;Meng Qingbing;Zhang Rui;Liu Liang;Xiao Hao;Tian Yingping(Department of Emergency,The Second Hospital of Hebei Medical University,Shijiazhuang 050000,Hebei,China)

机构地区：[1]河北医科大学第二医院急诊科,河北石家庄050000

出　　处：《中国中西医结合急救杂志》2023年第2期147-152,共6页Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care

摘　　要：目的 模拟革兰阳性球菌和革兰阴性杆菌混合感染,建立金黄色葡萄球菌肠毒素B(SEB)和脂多糖(LPS)复合诱导的小鼠脓毒症模型,观察血清炎症因子及组织蛋白表达水平的变化,探讨不同剂量胸腺肽-α1(Tα1)对脓毒症的免疫干预作用.方法 选择SPF级健康雄性BALb/c小鼠 90 只,按随机数字表法将小鼠分为对照组、Tα1 小剂量组和Tα1 大剂量组,每组 30 只.采用向小鼠腹腔注射 300 μg/kg SEB和1000 μg/kg LPS的方法复制小鼠复合感染脓毒症模型.于腹腔注射SEB2h后,给所有小鼠腹腔注射LPS 1000 μg/kg的同时Tα1 小剂量组小鼠腹部皮下注射Tα1100 μg/kg,Tα1 大剂量组小鼠腹部皮下注射Tα12000 μg/kg,对照组给予等量磷酸盐缓冲液(PBS).观察并记录各组小鼠制模后 6h的体征表现.于制模后2、4、6 h取血分离血清,采用酶联免疫吸附试验(ELISA)检测各组小鼠血清白细胞介素(IL-6,IL-10)、γ干扰素(IFN-γ)、肿瘤坏死因子-α(TNF-α)水平;处死动物后留取脾脏组织,光镜下观察脾脏组织的病理学改变,采用蛋白质免疫印迹试验(Western blotting)测定脾脏组织核转录因子-κB p65(NF-κB p65)的蛋白表达水平.结果 给药后 6 h,Tα1 小剂量组和Tα1 大剂量组小鼠精神活动、进食、寒战、呼吸、竖毛、大便等体征评分均较对照组明显降低[精神活动(分):1.20±0.42、1.10±0.31 比 2.70±0.48,进食(分):1.50±0.53、1.40±0.52比 2.60±0.70,寒战(分):1.20±0.42、1.30±0.48 比 2.30±0.48,呼吸(分):1.20±0.42、1.30±0.48 比 2.40±0.70,竖毛(分):1.40±0.52、1.20±0.42 比 2.60±0.52,大便(分):1.40±0.52、1.40±0.52 比 2.40±0.52,均P<0.05].随时间延长,各组IL-6、IL-10 逐渐降低,IL-10/IL-6 比值呈先降低后升高的趋势,IFN-γ呈先升高后降低的趋势;对照组TNF-α持续降低,脾脏组织NF-κB p65 的蛋白表达水平持续升高;Tα1 小剂量组和Tα1 大剂量组TNF-α呈先降低后升高趋势,脾脏组织NF-κB p65 的蛋白表Objective To simulate the combined infection of Gram positive coccus and Gram negative bacillus,and establish a mouse model of sepsis induced by Staphylococcus aureus enterotoxin B(SEB)and lipopolysaccharide(LPS),observe the expression levels of serum inflammatory factors and hiprotein,and explore the immune intervention effect of different doses of thymosinα1(Tα1)on sepsis.Methods Ninety SPF healthy male BALb/c mice were selected and divided into control group,Tα1 low-dose group,and Tα1 high-dose group according to random number table method,with 30 mice in each group.Mice were intraperitoneally injected with 300μg/kg SEB and 1000μg/kg LPS to reproduce a mouse model of complex sepsis.After 2 hours intraperitoneal injection of SEB,all mice were intraperitoneally injected with LPS 1000μg/kg,while mice in low-dose group were subcutaneously injected with Tαi100μg/kg,and mice in high-dose group were subcutaneously injected with Tα12000μg/kg,the control group was given the same amount of phosphate buffer salt solution(PBS).The physical manifestations of mice in each group were observed and recorded 6 hours after modeling.Blood samples were collected at 2,4,and 6 hours after modeling,and serum levels of interleukins(IL-6,IL-10),interferon-(IFN-γ),and tumor necrosis factor-α(TNF-α)were detected by enzyme-linked immunosorbent assay(ELISA).After death,spleen tissue was taken,and histopathological changes of spleen were observed under light microscope.Protein expression level of nuclear transcription factor-kB p65(NF-kB p65)in spleen tissue was determined by Western blotting.Results Six hours after administration,the scores of mental activity,feeding,shivering,breathing,hair erectomia,and stool in Tα1 low-dose group and Tα1 high-dose group were significantly lower than those in control group(mental activity:1.20±0.42,1.10±0.31 vs.2.70±0.48,feeding:1.50±0.53,1.40±0.52 vs.2.60±0.70,shivering:1.20±0.42,1.30±0.48 vs.2.30±0.48,breathing:1.20±0.42,1.30±0.48 vs.2.40±0.70,hair erectomia:1.40±0.52,1.20�

关 键 词：脓毒症 胸腺肽-Α1 免疫治疗 金黄色葡萄球菌肠毒素B 脂多糖 混合感染 

分 类 号：R459.7[医药卫生—急诊医学]