摘要
目的探讨酒精陛脂肪肝（ALD）肠源l生内毒素血症升高的始动因素，并观察四联活菌对ALD的干预效果。方法将50只雄性SD大鼠随机分为模型组（酒精灌胃组）、干预组（酒精和双歧四联活菌灌胃组）和对照组（等渗盐水灌胃组），喂养4、8周后分别取各组大鼠肝组织行HE染色观察肝脏病理学变化及电子显微镜下肠道黏膜变化、紧密连接occludin蛋白的表达变化，测定各组大鼠血清氨基转移酶、甘油三酯、内毒素的水平t取肠道新鲜粪便培养大肠杆菌。数据多组间比较采用单因素方差分析，差异有统计学意义者两两比较采用LDL—t检验。结果造模4周后，对照组、模型组及干预组的内毒素水平分别为（0．67±0．14）pg／ml、（4．42±1．28）pg／ml、（2．88±0．83）pg／ml，三组之间差异有统计学意义F=27．288，P〈0．01）；与对照组比较，模型组内毒素水平明显升高（P〈0．05）；与模型组比较，干预组内毒素水平明显降低（P〈0．05）。而三组之间ALT、AST、甘油三酯的差异并无统计学意义（P均〉0．05）。造模8周后，对照组、模型组及干预组的ALT和AST水平分别为（62．33±7．12）U／L和（90．50±10．67）U／L、（95．50±8．73）U／L和（130．00±14．91）U／L、（81．33±6．19）U／L和（110．33±7．26）U／L，差异均有统计学意义（F=18．051，P〈0．01lF=30．170，P〈0．01）；对照组、模型组及干预组的甘油三酯水平分别为（0．84土0．84）mmol／L、（1.40±0．17）mmol／L、（1．10±0．17）mmol／L，F=10。592，P〈0．01。对照组、模型组及干预组的大肠杆菌计数分别为（2．23±0．46）lg3／ml、（4．81±0．29）lg3／ml、（3．61±0．50）lg3／ml，差异有统计学意义（F=23．579，P〈0．01）；三组的内毒素水平分别为（0．52±0．21）pg／rnl、（12．46±2．61）pg／ml、（6．83±1．74）pg／ml，差异有统计学意义（F=30．731，JP〈0．01）。与对照组比较，模型组ALT、AST、甘油三酯、大肠杆菌计数及内毒素水平均明显增高（P值均〈0．05）；与模型组比较，干预组上述指标均明显降低p值均〈0．05）。造模8周后模型组比4周模型组小肠上皮细胞结构破坏更明显，细胞间隙增宽；干预组细胞连接结构比较清晰，细胞间隙稍增宽，造模8周后模型组occludin的表达强度明显下降，且分布不连续；而干预组occludin蛋白的表达水平则介于模型组与对照组之间。结论ALD早期存在肠道微生态紊乱并产生肠源性内毒素血症，肠道通透陛的增加可能是促使内毒素升高的始动因素；双歧四联活菌可能通过改变肠道菌群种类，上调肠道上皮黏膜occludin蛋白的表达，阻止内毒素通过紧密连接入血，从而有助于延缓ALD的进一步发展。
Objective To investigate the initial changes in the gut microenvironment that accompany intestinal endotoxemia related to alcoholic fatty liver disease （ALD） in order to explore the potential initiating factors and to observe the effect ofprobiotic therapy on these factors. Methods Fifty Sprague-Dawley male rats were randomly divided into an ALD model group （alcoholic intragastric administration）, an intervention group （ALD with probiotic intragastric administration）, and a control group （physiological saline intragastric administration）. Histological changes of the liver were evaluated using hematoxylin-eosin staining and light microscopy. Serum alanine aminotransferase （ALT）, aspartate aminotransferase （AST） and triglycerides （TG）, and plasma endotoxin and colibacillus were determined. The structural integrity of intestinal mucosa and tight junctions were observed by transmission electron microscopy. Occludin protein expression in intestinal epithelial cells was detected by immunohistochemistry. Results After four weeks, the three groups showed significant differences in the plasma endotoxin levels [control： （0.67± 0.14） pg/ml, model： （4.42± 1.28） pg/ ml, and intervention： （2.88 ± 0.83） pg/ml; F = 27.288, P = 0.000] and numbers of Escherichia coli [control： （2.31 ± 0.39） lg3/ml, model： （3.23 ± 0.41） lg3/ml, and intervention： （2.24 ± 0.44） lg3/ml; F= 10.692, P= 0.001]. The plasma endotoxin level and E. eoli number were significantly higher in the model group than in the control group and the intervention group （all P 〈 0.05）. The three groups showed no significant differences in the levels of ALT, AST, and TG at four weeks. After eight weeks, however, all three serum markers were significantly different between the three groups [ALT： control： （62.33 ~ 7.12） U/L, model： （95.50 ~ 8.73） U/L, and intervention： （81.33 ~ 6.19） U/L; F= 18.051, P = 0.000]; lAST： control： （90.50± 10.67） U/L, model： （130.00 ± 14.91） U/L, and intervention： （110.33± 7.26） U/L; F= 30.170, P = 0.000]; [TG： control： （0.84± 0.84） mmol/L, model： （1.40 ± 0.17） mmol/L, and intervention： （1.10 ± 0.17） mmol/L; F= 10.592, P = 0.001]. In addition, the three groups showed significant differences in E. coli number [control： （2.23± 0.46） lg3/ml, model： （4.81 ±0.29） lg3/ml, and intervention： （3.61 ±0.50） lg3/ml; F = 23.579, P = 0.000] and plasma endotoxin level [control： （0.52 ± 0.21） pg/ml, model： （12.46 ±2.61） pg/ml, intervention： （6.83±1.74） pg/ml; F= 30.731, P = 0.000]. The levels ofALT, AST, TG and endotoxin, and the number of E. coli were all significantly higher in the model group than in the control group and the intervention group （all P 〈 0.05）. Small intestinal epithelial cell structural failure was more apparent and intercellular gaps more broad after eight weeks than after four weeks for all three groups. However, the intervention group showed clearer cell connection structures and less extensive cell gap broadening than the model group at eight weeks. After eight weeks, the occludin protein had become significantly down-regulated and distributed in a non-continuous pattern in the model group, as compared with the control group. However, the occludin protein expression was higher in intervention group than in the model group. Conclusion Intestinal endotoxemia related to perturbations in the microenviroument occurs in the early phase of ALD, and the increased intestinal permeability appears to be the initial factor of elevated plasma endotoxin, which may lead to liver damage. Probiotlc therapy can reduced plasma endotoxin levels and postpone ALD progression by altering the composition of the gut microbiota and up-regulating expression of the occludin protein in intestinal epithelial cells.
出处
《中华肝脏病杂志》
CAS
CSCD
北大核心
2012年第11期848-852,共5页
Chinese Journal of Hepatology
基金
陕西省科技攻关科研基金[2007-k14-02（15）]
关键词
脂肪肝
酒精性
内毒素类
肠道微生态
紧密连接
Fatty liver, alcoholic
Endotoxins
Intestinal microecology
Tight junction
