重组旋毛虫53000抗原蛋白联合亚胺培南对多细菌感染脓毒症小鼠的保护作用  

Protective effects of recombinant trichinella spiralis-53 000 protein combining with imipenem on polymicrobial septic mice

作　　者：李凡[1] 陈志斌[1] 唐皓[1] 梁艳冰[1] 李振宇[1] 吴敬国[1] 曾丽金[1] 杨文[1] 胡旭初[2] 马中富[1] 

机构地区：[1]中山大学附属第一医院普内科,广东广州510080 [2]中山大学医学院寄生虫教研室,广东广州510080

出　　处：《中华危重病急救医学》2016年第5期433-438,共6页Chinese Critical Care Medicine

基　　金：广东省科技厅产业技术研究与开发资金计划项目（20128031800291）;广东省广州市科技攻关专项项目（201300000160）

摘　　要：目的观察重组旋毛虫53000抗原蛋白（rTsP53）联合亚胺培南（IMP）对脓毒症小鼠的保护作用，初步探讨其可能机制。方法按随机数字表法将雄性BALB／e小鼠分为5组，采用盲肠结扎穿孔术（CLP）构建多细菌感染小鼠脓毒症模型（CLP组），假手术（Sham）组仅开腹、关腹，不进行结扎。CLP＋IMP组、CLP＋rTsP53组、cLP＋IMP＋rTsP53组分别于术后6h起腹腔注射IMP20mg／kg＋0．1mL白蛋白、rTsP53蛋白6mg／kg＋0．1mL生理盐水（Ns）、IMP20mg／kg＋rTsP53蛋白6mg／kg；Sham组、CLP组则给予0．1mL白蛋白＋0．1mLNS；12h重复1次，至实验结束。各组取20只小鼠观察72h存活情况；并于术后0、6、12、24、36、48、72h各取3只小鼠血标本，采用酶联免疫吸附试验（ELISA）检测血清细胞因子水平，全血培养进行活菌菌落计数。24h处死小鼠取小肠组织，透射电镜下观察小肠黏膜上皮细胞超微结构。结果①cLP＋IMP＋rTsP53组小鼠存活率明显高于CLP组、CLP＋IMP组、CLP＋rTsP53组（85％比20％、55％、25％，均P〈0．05o②Sham组各时间点全血均未培养出细菌；各实验组术后6h活菌计数均显著升高，CLP组呈先升后降趋势，于24h达峰值（×10^6cfu／L：12．74±2．33）；CLP＋rTsP53组12h起显著高于CLP组，于36h达峰值（×10^6cfu／L：22．13±4．28）后逐渐下降；CLP＋IMP组、CLP＋IMP＋rTsP53组于6h达峰值（×10^6cfu／L：5．72±0．50、5．49±0．59）后呈逐渐下降趋势，12h起即显著低于CLP组。③各实验组术后6h起血清细胞因子水平均明显高于Sham组。CLP组肿瘤坏死因子-α（TNF-α）呈先升后降趋势，于36h达峰值（ng／L：1422：67±72．19），CLP＋IMP组、CLP＋rTsP53组、cIJP＋IMP＋rTsP53组达峰值时间提前至12h（nglL：1376．29＋44．67、929．36±40．42、809．61±22．61oCLP组和CLP＋IMP组白细胞介素-6（IL-6）于24h达峰值（ng/L：215．39±16．05、191．63±8．99），Objective To observe the protective effects of recombinant trichinella spiralis-53 000 protein （rTsP53 protein） combining with imipenem （IMP） on septic mice and their underlying mechanisms. Methods Male BALB/c mice were divided into five groups randomly. Cecal ligature and puncture （CLP） operation was used for building polymicrobial septic model （CLP group）. Mice in sham group were only subjected to laparoromy and abdominal closure without cecum ligation. At 6 hours post CLP, mice in CLP＋IMP, CLP＋rTsP53, and CLP＋IMP＋rTsP53 groups were injected intraperitoneally with IMP （20 mg/kg） ＋ 0.1 mL albumin, rTsP53 protein （6 mg/kg） ＋ 0.1 mL normal saline （NS）, IMP （20 mg/kg） ＋ rTsP53 protein （6 mg/kg） respectively, mice in sham group and CLP group were injected intraperitoneally with 0.1 mL albumin ＋ 0.1 mL NS, then these therapies were repeated every 12 hours until the experiment ended. Twenty mice were extracted randomly from each group for surveying 72-hour survival rate. At 0, 6, 12, 24, 36, 48, 72 hours post CLP, 3 mice in each group were collected and cytokines in serum were tested by enzyme-linked immunosorbent assay （ELISA）. Whole blood was cultured, then the numbers of bacteria colony-forming units （CFU） were counted. Mice were executed at 24 hours, then the epithelial cells ultrastructures of the mice small intestinal mucosa were observed by transmission electron microscope （TEM）. Results ① Compared with CLP, CLP＋IMP or CLP＋rTsP53 group, 72-hour survival rate of the mice in CLP＋IMP＋rTsP53 group was significantly elevated （85% vs. 20%, 55%, 25%, all P 〈 0.05）. ② No bacteria was found in cultured whole blood of mice in sham group at all time-points. At 6 hours post CLP operation, the number of bacterial clone of all experimental groups was rose significantly. The changed trend of bacterial number in CLP group was rising at the beginning, then declining, and the bacterial number reached the peak at 24 hours （×106 cfu/L： 12.74 ±2

关 键 词：重组旋毛虫53000抗原蛋白 亚胺培南 多细菌感染脓毒症 细胞因子 

分 类 号：R459.7[医药卫生—急诊医学]