丙戊酸钠对严重烫伤后肠屏障功能的保护作用及机制  

Protective effects of valproic acid on gut barrier function after major burn injury and its mechanism

作　　者：罗红敏[1] 胡森[2] 卞徽宁[1] 郑少逸[1] 熊兵[1] 黄志锋[1] 刘族安[1] 孙传伟[1] 马亮华[1] 李汉华[1] 喻文[3] 杜明华[4] 陈华德[1] 赖文[1] Luo Hongmin Hu Sen Bian Huining Zheng Shaoyi Xiong Bing Huang Zhifeng Liu Zu＇an Sun Chuanwei Ma Lianghua Li Hanhua Yu Wen Du Minghua Chen Huade Lai Wen(Department of Burns and Wound Repair Surgery, Guangdong General Hospital （Guangdong Academy of Medical Sciences）, Guangzhou 510080, Guangdong, China Laboratory of Shock and Organ Dysfunction, Burn Institute, First Hospital Affiliated to the PLA General Hospital, Beijing 100048, China Department of Critical Care Medicine, Sun Yat-sen Memorial Hospital, Guangzhou 510120, Guangdong, China Department of Emergency, Chinese PLA General Hospital, Beijing 100048. China)

机构地区：[1]广东省人民医院(广东省医学科学院)烧伤与创面修复外科,广东广州510080 [2]解放军总医院第一附属医院烧伤研究所休克与多器官障碍实验室,北京100048 [3]中山大学孙逸仙纪念医院重症医学科,广东广州510120 [4]解放军总医院急诊科,北京100853

出　　处：《中华危重病急救医学》2017年第3期221-227,共7页Chinese Critical Care Medicine

基　　金：国家自然科学基金（81301607）

摘　　要：目的探讨丙戊酸钠（VPA）对致死性烫伤大鼠肠屏障的保护作用及其机制。方法清洁级雄性SD大鼠40只，按随机数字表法分为假烫（Sham）＋生理盐水（NS）组、Sham＋VPA组、烫伤（Scald）＋NS组、Scald＋VPA组4组，每组10只。将大鼠浸入80℃水中建立55％总体表面积（TBSA）Ⅲ度烫伤休克大鼠模型，Sham组大鼠浸入37℃水中相同时间；烫伤后即刻皮下注射300mg／kg VPA或NS 0.25mL。于伤后2h、6h处死大鼠取腹主动脉血和回肠组织，采用酶联免疫吸附试验（ELISA）检测肠组织血管内皮生长因子（VEGF）水平；检测异硫氰酸荧光素标记的葡聚糖（FITC-dextran）水平反映肠上皮屏障的通透性变化；采用Chiu肠屏障损伤评分评价肠黏膜组织形态学变化；采用免疫荧光和蛋白质免疫印迹试验（Western Blot）检测肠组织乙酰化组蛋白3第9位赖氨酸（Ac-H3K9）水平以及缺氧诱导因子1α（HIF-1α）、带状闭合蛋白1（ZO-1）、肌球蛋白轻链激酶（MLCK）的蛋白表达。结果与Sham＋NS组相比，Scald＋NS组大鼠伤后2h即表现出肠黏膜组织损伤，肠黏膜通透性增加，肠组织组蛋白乙酰化下降，ZO-1降解增加，HIF-1α、VEGF及MCLK蛋白表达上调，ZO-1蛋白表达下调；且上述变化在伤后6h更为明显。VPA可明显逆转55％TBSAⅢ度烫伤造成的肠黏膜损伤表现。与Scald＋NS组相比，Scald＋VPA组伤后2h上述逆转效果尚不明显；而6h肠黏膜损伤明显减轻[Chiu评分（分）：2．03±0．27比3．12±0．15]，肠黏膜通透性明显降低[FITC-dextran（μg/L）：709±76比1138±75]，组蛋白乙酰化增加[Ac-H3K9（灰度值）：1．55±0．12比0．48±0．12]，ZO-1降解明显抑制（灰度值：0．69±0．12比0．43±0．16），VEGF水平和MLCK蛋白表达明显下调[VEGF（ng／mg）：51．7±3．7比71．2±4．3，MLCK（灰度值）：1．98±0．20比2．80±0．24]，HIF-1α蛋白在伤后2h和6h均明显下调（�Objective To investigate the potential protective effects of valproic acid （VPA） on gut barrier function after major burn injury in rats and its mechanism. Methods Forty male Sprague-Dawley （SD） rats were divided into sham ＋ normal saline （NS）, sham ＋ VPA, scald ＋ NS, and scald ＋ VPA groups, with 10 rats in each group. Rat with 55% total body surface area （TBSA） third-degree severe-burns model was reproduced by immersing into 80℃ water, and the rats in sham groups were given sham-burns by immersing into 37℃ water. The rats after severe- burns were immediately treated with 0.25 mL of 300 mg/kg VPA or NS by subcutaneous injection. Rats were sacrificed at 2 hours and 6 hours after injury, and abdominal aortic blood and ileal tissue were harvested. The levels of vascular endothelial growth factor （VEGF） were determined by enzyme-linked immunosorbent assay （ELISA）. The intestinal permeability was evaluated by fluorescein isothiocyanate-dextran （FITC-dextran） determination. The histomorphological changes in gut barrier were evaluated by Chiu grading system. Levels of acetylated lysine at the ninth position of histone 3 protein （Ac-H3K9）, hypoxia-inducible factor 1α （HIF-1α）, zona occludens 1 （ZO-1） and myosin light chain kinase （MLCK） were determined by immunofluorescence staining and Western Blot. Results Compared with sham ＋ NS group, rats in scald ＋ NS group showed intestinal mucosal damage 2 hours after burn injury, as well as increased mucosal permeability, protein expression levels of HIF-1α, VEGF, MLCK, and lowered levels of AC-H3K9 and ZO-1. These changes were much more prominent at 6 hours after injury. VPA treatment significantly attenuated the burn-induced intestinal damage. Compared with scald ＋ NS group, the protective effects in scald ＋ VPA group was not evident at 2 hours after injury; however, intestinal damage was much less severe at 6 hours after injury （Chiu score： 2.03 ± 0.27 vs. 3.12 ± 0.15）, intestinal permeability was sign

关 键 词：烧伤 肠屏障 组蛋白去乙酰化酶抑制剂 丙戊酸钠 缺氧诱导因子1Α 

分 类 号：R644[医药卫生—外科学]