动物营养学报    2022, Vol. 34 Issue (6): 4003-4012    PDF    
三丁酸甘油酯对产肠毒素大肠杆菌K88诱发的小鼠肠道损伤的缓解作用
庞丽丽1,2 , 朱荣生1 *, 王怀中1 , 王建才1 , 齐静1 , 黄保华1 , 呼红梅1     
1. 山东省农业科学院畜牧兽医研究所, 济南 250100;
2. 国家北京药物安全评价研究中心, 北京 100850
摘要: 本试验旨在研究三丁酸甘油酯(TB)对产肠毒素大肠杆菌K88(ETEC K88)诱导的小鼠肠道损伤的缓解作用。试验1:选用(20±2) g的BALB/c小鼠50只, 随机分为5组, 每组10只。各组均饲喂基础饲粮, 在第4天对照组腹腔注射0.5 mL无菌磷酸盐缓冲液(PBS), 试验组腹腔分别注射浓度为5.5×108、4.9×107、4.0×106、6.5×105 CFU/mL的ETEC K88悬液0.5 mL, 连续观察3 d临床症状, 统计腹泻率和死亡率, 剖检、观察肠道病理变化, 确定ETEC K88最佳造模剂量。试验2:选择(20±2) g的BALB/c小鼠105只, 随机分为7组, 每组3个重复, 每个重复5只。空白组、阴性对照组饲喂基础饲粮, 阳性对照组饲喂添加0.16%金霉素的饲粮, 试验1、2、3、4组分别饲喂添加0.05%、0.10%、0.20%、0.30%TB的饲粮。连续饲喂14 d, 在第15天空白组小鼠腹腔注射0.5 mL无菌PBS, 阴性对照组、阳性对照组和试验组小鼠按照试验1中确定的最佳造模剂量腹腔注射ETEC K88悬液0.5 mL, 继续观察3 d, 采血、剖检, 检测血清中二胺氧化酶(DAO)活性与内毒素(ET)、C-反应蛋白(CRP)和分泌型免疫球蛋白A(SIgA)含量, 制作空肠组织切片, 观察肠道组织形态变化。结果显示: 小鼠腹腔注射不同浓度ETEC K88悬液后出现不同程度的腹泻和死亡情况, 根据腹泻率和死亡率, 确定4.9×107 CFU/mL为ETEC K88建立小鼠腹泻模型的最佳剂量。阳性对照组和试验3、4组的平均日增重与空白对照组差异不显著(P < 0.05), 但显著高于阴性对照组(P>0.05)。小鼠的腹泻率表现为阴性对照组(53.33%)>试验1组(33.33%)>阳性对照组(26.67%)=试验2组(26.67%)>试验3组(20.00%)=试验4组(20.00%)>空白对照组(无腹泻发生)。肠道损伤程度: 阴性对照组>试验1组>阳性对照组>试验2组>空白对照组>试验4组>试验3组。与阴性对照组相比, 各试验组以及阳性对照组小鼠血清CRP、ET含量和DAO活性均有不同程度降低, 以试验3组降低幅度最大, 其次为试验4组。与阳性对照组相比, 试验3、4组小鼠血清CRP、ET含量和DAO活性均有不同程度降低, 但差异不显著(P>0.05)。与阴性对照组相比, 试验3、4组小鼠血清SIgA含量分别增加了169.00%(P < 0.05)、114.33%(P < 0.05)。与阳性对照组相比, 试验3、4组小鼠血清SIgA含量分别增加了72.81%(P < 0.05)和37.69%(P < 0.05)。由此得出, 在小鼠基础饲粮中添加0.20%或0.30%的TB可以对抗大肠杆菌感染, 缓解ETEC K88诱导的肠道损伤, 且效果与添加0.16%的金霉素相当。
关键词: 三丁酸甘油酯    产肠毒素大肠杆菌    小鼠腹泻模型    肠道损伤    
Alleviation Effects of Triglyceride on Intestinal Injury of Mice Induced by Enterotoxigenic Escherichia coli K88
PANG Lili1,2 , ZHU Rongsheng1 *, WANG Huaizhong1 , WANG Jiancai1 , QI Jing1 , HUANG Baohua1 , HU Hongmei1     
1. Institute of Animal Husbandry and Veterinary Medicine, Shandong Academy of Agricultural Sciences, Jinan 250100, China;
2. National Beijing Center for Drug Safety Evaluation and Research, Beijing 100850, China
Abstract: This experiment was conducted to study the alleviation effect of triglyceride (TB) on intestinal injury of mice induced by enterotoxigenic Escherichia coli K88 (ETEC K88). Experiment 1: fifty BALB/c mice with the body weight of (20±2) g were selected and randomly divided into 5 groups with 10 mice per group. All mice fed the same basal diet, and the mice in control group were intraperitoneal injection of 0.5 mL aseptic phosphate buffer solution (PBS), while those in experimental groups were intraperitoneal injection of 0.5 mL ETEC K88 suspension with the concentrations of 5.5×108, 4.9×107, 4.0×106 and 6.5×105 CFU/mL, respectively. The clinical symptoms were observed and recorded within three days, calculated the diarrhea rate and mortality. Then, necropsied the mice to observe their intestinal pathological changes to determine the best dose of ETEC K88 for modeling. Experiment 2: one hundred and five mice with the body weight of (20±2) g were selected and randomly divided into 7 groups with 3 replicates per group and 5 mice per replicate. The negative control group intraperitoneal injection of the ETEC K88 suspension (best dose determined in experiment 1, the same as below) after fed the basal diet for 14 days, the blank control group intraperitoneal injection of aseptic PBS after fed the basal diet for 14 days, the positive control group intraperitoneal injection of ETEC K88 suspension after fed the basal diet added 0.16% chlortetracycline for 14 days, and the four experimental groups intraperitoneal injection of ETEC K88 suspension after fed the basal diet added 0.05%, 0.10%, 0.20% and 0.30% TB for 14 days. After observation for 3 days, collected blood and necropsied the mice, then detected the changes of diamine oxidase (DAO) activity, endotoxin (ET), C-reactive protein (CRP), and secretory immunoglobulin A (SIgA) contents in serum and made intestinal pathology sections to observe the intestinal histological changes. The results showed that, after intraperitoneal injection of different concentrations of ETEC K88 suspension, the mice developed diarrhea and death of varying degrees. The best dose of ETEC K88 to build mice diarrhea model was determined according to the diarrhea rate and mortality, it was 4.9×107 CFU/mL. The average daily gain of positive control group and experimental groups 1 and 2 had no significant difference compared with the blank control group (P < 0.05), but significantly higher than that of negative control group (P>0.05) The diarrhea rate of mice was negative control group (53.33%)>experimental group 1 (33.33%)>positive control group (26.67%)=experimental group 2 (26.67%)>experimental group 3(20.00%)=experimental group 4(20.00%)>blank control group (no diarrhea). Degree of intestinal injury manifested as: negative control group>experimental group 1>positive control group>experimental group 2>blank control group>experimental group 4>experimental group 3. Compared with the negative control group, the serum CRP and ET content and DAO activity of experimental groups 3, 4 and positive control group were reduced in different degrees, and experimental group 3 had the biggest reduction, followed by experimental group 4. the serum CRP and ET content and DAO activity of experimental groups 3, 4 were reduced in different degrees compared with the positive control group, but there were no significant differences (P>0.05). Serum SIgA content of experimental groups 3, 4 were increased by 169.00% (P < 0.05) and 114.33% (P < 0.05) compared with the negative control group, respectively, and they were increased by 72.81% (P < 0.05) and 37.69% (P < 0.05) compared with the positive control group, respectively. In conclusion, adding 0.20% or 0.30% TB to the basal diet of mice can combat Escherichia coli infection, alleviate intestinal injury induced by ETEC K88, and the effect can be comparable to 0.16% chlortetracycline.
Key words: tributyrin    enterotoxigenic Escherichia coli    mice diarrhea model    intestinal injury    

仔猪腹泻是生产中典型的多因素疾病,也是造成仔猪死亡的主要原因。仔猪腹泻的病因和流行性病学非常复杂,产肠毒素大肠杆菌(enterotoxigenic Escherichia coli,ETEC)是引起仔猪腹泻最常见的食源性流行病原体[1]。ETEC引起腹泻主要与菌毛黏附素和肠毒素有关,它们促进病原菌黏附于肠上皮细胞,导致机体电解质紊乱、酸碱失衡和肠道菌群失调,损伤肠道正常形态,引发炎症反应[2-3]。据统计,ETEC感染引起的仔猪腹泻在国内约占35%,在美国等其他国家约占45%,是导致生猪养殖业损失的重要因素[4-5]。目前抗生素疗法是治疗仔猪大肠杆菌性腹泻的主要措施,但是易反复和继发感染,而且易导致动物肠道菌群失衡和肠壁损伤[6]。三丁酸甘油酯(tributyrin,TB)作为丁酸前体物,因其为肠道上皮细胞供能,刺激肠上皮细胞增殖等特点而受到关注[7],其可改善肠道黏膜的营养状态,修复抗生素、乙酸等因素导致的肠道损伤[6, 8-9]。研究表明,饲粮中添加0.1%TB可促进肠道发育,提高营养物质的消化吸收[10],维持肠道菌群平衡[11-12],提高免疫力,促进黏蛋白分泌,进而提高畜禽生产性能[13-18];饲粮添加0.1%TB可显著减缓乙酸和抗生素(甲硝唑、硫酸新霉素、万古霉素,每日分别给药1 g、500 mg、1 mg,连续7 d)诱发的肠道损伤[11-12];给小鼠口服或灌肠TB,可减轻硫酸葡聚糖钠(dextran sulphate sodium,DSS)诱导的结肠黏膜损伤和炎症[19]。金霉素是治疗细菌疾病的常用抗生素,饲粮中添加75 mg/kg金霉素可显著降低仔猪腹泻,提高生长性能[20]。目前,可通过腹腔注射、鼻饲、灌服大肠杆菌的方法建立稳定的小鼠腹泻模型。ETEC作为外源菌,感染后引起动物腹泻、诱导肠道炎症发生,腹泻模型已成为研究外源菌感染诱发的肠炎、病原菌与宿主免疫系统相互关系、微生态制剂、益生菌等治疗肠炎机制的理想模型[21]。目前对于TB是否能够修复ETEC诱发的肠道损伤未见报道,为此,本试验以ETEC K88诱导建立的小鼠腹泻模型为载体,研究TB对ETEC K88诱发的小鼠肠道损伤的影响,为通过营养手段改善大肠杆菌诱发的肠道损伤,促进肠道屏障完善,减少腹泻发生提供理论依据。

1 材料与方法 1.1 试验材料 1.1.1 试验动物

选取体重相近的健康BCLB/c小鼠155只,体重为(20±2) g,由济南新建生物技术有限公司提供。小鼠基础饲粮由青岛可奈尔饲料有限公司提供。试验期间,试验小鼠饲养于相对湿度在30%~40%的清洁动物笼中,自由采食、饮水。垫料高压灭菌,每天更换1次。

1.1.2 ETEC K88

ETEC K88标准株(CVCC196)购自中国菌种保存中心。

1.2 试验设计 1.2.1 ETEC K88诱发小鼠腹泻模型的构建

选择体重相近的BALB/c小鼠50只,随机分为5组,每组10只。各组均饲喂基础饲粮,在第4天对照组腹腔注射无菌磷酸盐缓冲液(PBS)0.5 mL,试验1、2、3、4组分别腹腔注射浓度为5.5×108、4.9×107、4.0×106、6.5×105 CFU/mL的ETEC K88悬液0.5 mL。连续观察3 d,记录小鼠的临床症状、腹泻数量和死亡数量。

1.2.2 TB对ETEC K88诱发的肠道损伤的缓解作用

选择体重相近的BALB/c小鼠105只,随机分为7组,每组3个重复,每个重复5只。空白对照组、阴性对照组饲喂基础饲粮,阳性对照组饲喂添加0.16%金霉素的基础饲粮,试验1、2、3、4组分别饲喂添加0.05%、0.10%、0.20%、0.30%TB的基础饲粮,连续饲喂14 d。在第15天,空白组腹腔注射无菌PBS 0.5 mL,阴性对照组、阳性对照组和试验1、2、3、4组按照1.2.1中确定的造模剂量腹腔注射ETEC K88悬液0.5 mL。连续观察3 d,记录小鼠的临床症状、腹泻和死亡数量,在注射后第4天采血并剖检小鼠,观察肠道病变。试验小鼠分组和处理见表 1

表 1 试验小鼠分组及饲喂方案 Table 1 Grouping and dosage regimen of experimental mice
1.3 检测指标

腹泻模型构建过程中观察小鼠精神状况、行动力、被毛、排便情况,记录腹泻、死亡数量。使用紫外分光光度计测定菌悬液在560 nm出的吸光度(OD560 nm),确定菌悬液细菌浓度(OD560 nm=0.8时,细菌浓度约为108 CFU/mL)。

在TB对ETEC K88诱发的肠道损伤的缓解作用试验中连续观察小鼠精神状况、行动力、被毛状况,记录小鼠体重、采食情况。参考Swaggerty等[22]的方法对肠道病变进行评分并制作肠道组织病理切片,采用酶联免疫吸附测定(ELISA)方法测定血清中二胺氧化酶(diamine oxidase,DAO)活性与内毒素(endotoxin,ET)、C-反应蛋白(C-reactive protein,CRP)和分泌型免疫球蛋白A(secretory immunoglobulin A,SIgA)含量。

1.4 数据统计分析

应用Graphpad prism 6.0软件对数据进行方差分析,采用LSD法进行多重比较。P < 0.05为差异显著,P < 0.01为差异极显著,结果均以“平均值±标准误(SE)”表示。

2 结果与分析 2.1 ETEC K88诱发小鼠腹泻模型的建立

对照组小鼠腹腔注射无菌PBS后2 h恢复行动能力,开始采食饮水,无腹泻和死亡。试验组小鼠腹腔注射ETEC K88悬液后出现不同程度的腹泻,而且被毛逆立粗乱、精神沉郁、趴窝不动、反应迟缓。试验1、2、3、4组小鼠腹腔注射ETEC K88悬液浓度分别为5.5×108、4.9×107、4.0×106、6.5×105 CFU/mL,小鼠的腹泻率分别为100%、80%、20%、0%,死亡率分别为90%、20%、0%、0%,具体见表 2。拟合小鼠腹泻率和死亡率曲线,根据改良寇氏法计算得知半数致死剂量(LD50)为4.9×107 CFU/mL,因此选择腹腔注射4.9×107 CFU/mL ETEC K88建立小鼠腹泻模型,研究TB对ETEC K88诱发的肠道损伤的缓解作用。

表 2 产肠毒大肠杆菌K88对小鼠腹泻率和死亡率的影响 Table 2 Effects of ETEC K88 on diarrhea and mortality in mice
2.2 TB对ETEC K88诱发的小鼠肠道损伤的缓解作用 2.2.1 TB对小鼠增重的影响

表 3可知,各组试验小鼠的初始体重相近,差异不显著(P>0.05)。与阴性对照组相比,阳性对照组和试验3、4组的平均日增重显著增加(P < 0.05),分别提高了27.78%、25.00%和34.95%。阳性对照组和试验3、4组的平均日增重均显著高于试验1、2组(P < 0.05),提高了17.65%~27.85%。

表 3 TB对小鼠增重的影响 Table 3 Effects of TB on weight gain of mice
2.2.2 TB对小鼠腹泻率和死亡率的影响

表 4可知,阴性对照组小鼠的腹泻率和死亡率最高,分别达到53.33%和20.00%,其他各组均无死亡。其余各组中,试验1组腹泻率最高,达到33.33%,其次为阳性对照组和试验2组,腹泻率均为26.67%,试验3、4组腹泻率均为20.00%,空白对照组无腹泻发生。由此可见,饲粮中添加TB和金霉素对ETEC K88诱发的小鼠腹泻和死亡均有较好的预防和缓解作用,其中饲粮中添加0.20%、0.30%TB时效果明显,优于添加金霉素。

表 4 TB对小鼠腹泻率和死亡率的影响 Table 4 Effects of TB on diarrhea rate and mortality of mice
2.2.3 TB对小鼠肠道组织病变的影响

剖检正常、腹泻和死亡小鼠,观察肠道病理变化,并进行评估。死亡小鼠的肠道组织损伤比腹泻小鼠严重,未发病小鼠肠道组织未见明显病变。死亡小鼠空肠达到4级病变,大面积广泛性出血;十二指肠2~3级病变,不同程度肿胀,分布出血带;盲肠直肠1级病变,有肉眼可见出血点。腹泻小鼠空肠3级病变,肉眼可见出血点,肠道有水样填充物;十二指肠1~2级病变,肠壁充盈变薄,偶见肠腔有气体;盲肠直肠0级病变,眼观无病变。由此可见,腹腔注射的ETEC K88主要侵袭肠道近端,空肠病变最严重,为此本试验进一步对腹泻小鼠(空白对照组为正常小鼠)空肠进行HE染色处理,进一步观察其形态和病理变化。

2.2.4 TB对小鼠空肠组织形态的影响

图 1可见,各组小鼠空肠损伤程度差别明显,就肠道组织完整性来看,损伤程度由大到小的顺序:阴性对照组>试验1组>阳性对照组>试验2组>空白对照组>试验4组>试验3组。与空白对照组相比,阴性对照组小鼠空肠黏膜损伤严重,肠绒毛断裂,杯状细胞、浆细胞大量浸润。与阴性对照组相比,阳性对照组和试验1、2组小鼠空肠黏膜组织较完整,肠绒毛边缘不清晰,绒毛形态较完整,可见隐窝,上皮细胞排列松散;试验3、4组小鼠肠组织结构清晰,绒毛边缘清晰、完整、排列整齐,上皮细胞排列紧密。由此可见,饲粮中添加0.20%、0.30%TB可有效减轻ETEC K88对小鼠肠道的损伤。

A:空白对照组blank control group;B:阴性对照组negative control group;C:阳性对照组positive control group;D:试验1组experimental group 1;E:试验2组experimental group 2;F:试验3组experimental group 3;G:试验4组experimental group 4。 图 1 空肠组织切片 Fig. 1 Jejunum tissue section (100×)
2.2.5 TB对小鼠血清炎症相关指标的影响

表 5可知,与空白对照组相比,其他各组小鼠腹腔注射ETEC K88后血清CRP、ET含量与DAO活性均有不同程度增加,以阴性对照组增加幅度最大,分别增加了128.42%(P < 0.05)、85.88%(P < 0.05)和298.79%(P < 0.01),以试验3组增加幅度最小,分别增加了65.47%(P>0.05)、5.65%(P>0.05)和101.61%(P < 0.01)。与阴性对照组相比,其他各组小鼠血清CRP、ET含量和DAO活性均有不同程度降低,以试验3组降低幅度最大,分别降低了27.56%(P>0.05)、43.16%(P < 0.05)和49.44%(P < 0.01),其次为试验4组,分别降低了23.62%(P>0.05)、39.67%(P < 0.05)和46.40%(P < 0.01)。与阳性对照组相比,试验3、4组小鼠血清CRP、ET含量和DAO活性均有不同程度降低,但差异不显著(P>0.05)。试验1、2、3、4组间小鼠血清CRP、ET含量和DAO活性相近,以试验3组最低,其次为试验4组,试验2组较高,试验1组最高。试验3组小鼠血清ET含量显著低于试验1组(P < 0.05),降低了39.19%。与空白对照组相比,试验1、2、4组及阳性对照组和阴性对照组小鼠血清SIgA含量均有不同程度降低,分别降低了71.80%(P < 0.01)、61.33%(P < 0.05)、44.38%(P < 0.05)、68.25%(P < 0.05)、74.05%(P < 0.01),试验3组只在数值上降低了30.19%,差异不显著(P>0.05)。与阴性对照组相比,试验3、4组小鼠血清SIgA含量显著增加(P < 0.05),分别增加了169.00%、114.33%。与阳性对照组相比,试验3、4组小鼠血清SIgA含量显著增加(P < 0.05),分别增加了72.81%、37.69%。

表 5 TB对小鼠血清炎症相关指标的影响 Table 5 Effects of TB on serum inflammation-related indexes of mice
3 讨论

本课题组前期研究发现,在仔猪饲粮中添加0.10%~0.15%的TB可替代抗生素等药物添加剂,同时仔猪的生长性能、肠组织发育和免疫功能得到改善,添加0.20%~0.25%的TB不仅可替代抗生素等药物添加剂,而且显著提升仔猪的生长性能、养分消化率和健康水平,显著增强肠组织功能[23]。本试验主要研究TB对ETEC K88诱发的肠道损伤的缓解作用,为其在仔猪上的应用提供借鉴。

本研究发现,给小鼠饲喂添加不同水平TB的饲粮14 d后,腹腔注射ETEC K88悬液后小鼠腹泻率和死亡率均明显低于阴性对照组小鼠,其中添加0.10%TB的试验2组与阳性对照组的腹泻率和死亡率相当,添加0.20%、0.30%TB的试验2、3组的腹泻率低于阳性对照组,表明饲粮中添加TB可有效缓解ETEC K88引起的小鼠腹泻,且添加0.20%或0.30%TB时效果优于添加0.16%金霉素。腹腔注射ETEC K88后小鼠的平均日增重减少17.24%,而饲喂TB小鼠的平均日增重均有不同程度增加,其中添加0.20%和0.30%TB时平均日增重显著增加,效果与添加0.16%金霉素相当。由此可见,饲粮添加TB可缓解ETEC K88对小鼠增重的负面影响,与Hou等[9]和Wang等[24]的研究结果一致。这可能与TB可有效改善小鼠的肠道消化吸收能力和健康状态有关。

肠道黏膜不仅是营养物质消化吸收的场所,还是抵御外界病原体进入机体的屏障。本研究发现,腹腔注射的ETEC K88主要侵袭肠道近端,以空肠病变最严重,表现为肠绒毛断裂,肠腺消失,杯状细胞、浆细胞大量浸润。饲粮中添加0.20%、0.30%TB可有效缓解ETEC K88诱导产生的小鼠肠道损伤,保护肠组织结构完整,使肠绒毛边缘清晰、完整、排列整齐,隐窝清晰,肠腺明显,上皮细胞排列紧密。这与前人研究结果[25-26]一致。饲粮添加TB可改善肠炎肉鸡的肠道黏膜屏障功能,提高营养物质吸收能力[25-26],而且TB可提高仔猪肠黏膜表皮生长因子受体(EGFR)基因的表达,通过激活EGFR信号缓解乙酸诱发的肠道损伤,使平均日增重提高13.81%[27]。抗生素可降低肠道中紧密连接蛋白的表达,以及与离子交换体、丁酸转运相关的基因和蛋白的表达,而饲粮添加TB可显著减少这些损失[6]

SIgA是由肠黏膜上皮细胞分泌的一种关键性抗炎抗体[28]。肠道黏膜分泌SIgA是保护肠道感染的主要防御机制之一,因此被用作细菌致病性的标志指标[29]。当外界或内部抗原刺激时,肠黏膜分泌SIgA和细胞因子等物质维持肠上皮的稳态[26]。DAO是一种存在于哺乳动物小肠黏膜和肠绒毛上皮细胞中催化二胺的细胞内酶,通过调节细胞内的离子平衡、促进细胞修复保护肠道黏膜。肠道黏膜屏障功能衰竭时或肠黏膜细胞坏死时血清DAO活性升高,因此DAO活性可反映肠道损伤状态[27, 30]。肠道屏障功能受损后ET转移入血,造成血液中ET含量升高,研究表明动物大肠杆菌感染后血清ET含量升高明显[31-32]。CRP是在机体受到感染或组织损伤时血浆中一些急剧上升的蛋白质(急性蛋白)[33],细菌感染时血清CRP的含量明显升高至中等度,阳性率可达90%以上,而病毒等感染时,血清这个CRP含量多为正常或轻度升高,因此CRP含量常被用来鉴别诊断细菌感染与非细菌感染[34-35]。试验小鼠腹腔注射ETEC K88后,血清SIgA含量显著降低,DAO活性显著增加,ET、CRP含量显著升高,符合细菌感染肠道损伤后导致的炎症相关指标变化;且试验结果表明饲粮添加金霉素和TB均可有效减缓这些变化,综合比较发现饲粮添加0.20%或0.30% TB比添加0.16%金霉素的作用效果更明显。由此可见,饲粮中添加0.20%或0.30%的TB可有效缓解由ETEC K88诱导产生的小鼠肠道损伤,降低肠道的通透性,保护肠道黏膜。

4 结论

① 腹腔注射4.9×107 CFU/mL ETEC K88悬液0.5 mL可以成功构建小鼠腹泻模型。

② 在小鼠基础饲粮中添加0.20%或0.30%的TB可缓解由ETEC K88诱导产生的肠道损伤,且效果优于添加0.16%的金霉素。

参考文献
[1]
LIU G, REN W K, FANG J, et al. L-glutamine and L-arginine protect against enterotoxigenic Escherichia coli infection via intestinal innate immunity in mice[J]. Amino Acids, 2017, 49(12): 1945-1954. DOI:10.1007/s00726-017-2410-9
[2]
BIN P, TANG Z Y, LIU S J, et al. Intestinal microbiota mediates enterotoxigenic Escherichia coli-induced diarrhea in piglets[J]. BMC Veterinary Research, 2018, 14(1): 385. DOI:10.1186/s12917-018-1704-9
[3]
YANG K M, JIANG Z Y, ZHENG C T, et al. Effect of Lactobacillus plantarum on diarrhea and intestinal barrier function of young piglets challenged with enterotoxigenic Escherichia coli K88[J]. Journal of Animal Science, 2014, 92(4): 1496-1503. DOI:10.2527/jas.2013-6619
[4]
LU T, MOXLEY R A, ZHANG W P. Application of a novel epitope- and structure-based vaccinology-assisted fimbria-toxin multiepitope fusion antigen of enterotoxigenic Escherichia coli for development of multivalent vaccines against porcine postweaning diarrhea[J]. Applied and Environmental Microbiology, 2020, 86(24): e00274-20.
[5]
ZHANG W P, ZHAO M J, RUESCH L, et al. Prevalence of virulence genes in Escherichia coli strains recently isolated from young pigs with diarrhea in the US[J]. Veterinary Microbiology, 2007, 123(1/2/3): 145-152.
[6]
CRESCI G, NAGY L E, GANAPATHY V. Lactobacillus GG and tributyrin supplementation reduce antibiotic-induced intestinal injury[J]. JPEN.Journal of Parenteral and Enteral Nutrition, 2013, 37(6): 763-774. DOI:10.1177/0148607113486809
[7]
王萌, 黄铁军, 张勇. 三丁酸甘油酯对断奶仔猪生长性能及肠道健康的调控[J]. 饲料工业, 2016, 37(12): 56-58.
WANG M, HUANG T J, ZHANG Y. Regulation of tributyrin on growth performance and intestinal health in weaning piglets[J]. Feed Industry, 2016, 37(12): 56-58 (in Chinese).
[8]
PIVA A, GRILLI E, FABBRI L, et al. Intestinal metabolism of weaned piglets fed a typical United States or European diet with or without supplementation of tributyrin and lactitol[J]. Journal of Animal Science, 2008, 86(11): 2952-2961. DOI:10.2527/jas.2007-0402
[9]
HOU Y Q, WANG L, YI D, et al. Dietary supplementation with tributyrin alleviates intestinal injury in piglets challenged with intrarectal administration of acetic acid[J]. British Journal of Nutrition, 2014, 111(10): 1748-1758. DOI:10.1017/S0007114514000038
[10]
李丹丹, 宇晓军, 王群, 等. 三丁酸甘油酯对育成期母貂生长性能、营养物质表观消化率及十二指肠消化酶活性的影响[J]. 动物营养学报, 2021, 33(6): 3461-3468.
LI D D, YU X J, WANG Q, et al. Effects of tributyrin on growth performance, nutrient apparent digestibilities and duodenum digestive enzyme activities of growing female minks[J]. Chinese Journal of Animal Nutrition, 2021, 33(6): 3461-3468 (in Chinese). DOI:10.3969/j.issn.1006-267x.2021.06.047
[11]
MIRAGOLI F, PATRONE V, PRANDINI A, et al. Implications of tributyrin on gut microbiota shifts related to performances of weaning piglets[J]. Microorganisms, 2021, 9(3): 584. DOI:10.3390/microorganisms9030584
[12]
梁琦, 左刚. 三丁酸甘油酯的生理功能及其在仔猪中的应用[J]. 猪业科学, 2020, 37(5): 46-49.
LIANG Q, ZUO G. Physiological functions of tributyrin and its application in piglets[J]. Swine Industry Science, 2020, 37(5): 46-49 (in Chinese). DOI:10.3969/j.issn.1673-5358.2020.05.013
[13]
陈思, 杜红方, 陈书琴, 等. 博落回和三丁酸甘油酯对黄羽肉鸡生长性能及肠道形态的影响[J]. 中国畜牧杂志, 2021, 57(7): 224-229.
CHEN S, DU H F, CHEN S Q, et al. Effects of Boluohui and tributyrin on growth performance and intestinal morphology of yellow-feather broilers[J]. Chinese Journal of Animal Science, 2021, 57(7): 224-229 (in Chinese).
[14]
彭宇, 柴毛毛, 崔细鹏, 等. 丁酸类添加剂及其与合生元组合对肉鸡生长性能和肠道健康的影响[J]. 动物营养学报, 2020, 32(11): 5145-5157.
PENG Y, CHAI M M, CUI X P, et al. Effects of butyric acids additives alone or combination with synbiotics on growth performance and gut health of broilers[J]. Chinese Journal of Animal Nutrition, 2020, 32(11): 5145-5157 (in Chinese). DOI:10.3969/j.issn.1006-267x.2020.11.020
[15]
张文华. 丁酸甘油酯对猪健康的作用[J]. 国外畜牧学(猪与禽), 2018, 38(9): 18-19.
ZHANG W H. Glyerides of butyric acid for pig health[J]. Animal Science Abroad (Pigs and Poultry), 2018, 38(9): 18-19 (in Chinese). DOI:10.3969/j.issn.1001-0769.2018.09.005
[16]
LIU Z Y, YANG H L, YAN Y Y, et al. Supplementation of tributyrin, alone and in combination with fructooligosaccharide in high soybean meal diets for shrimp (Litopenaeus vannamei): effects on growth, innate immunity and intestinal morphology[J]. Aquaculture Nutrition, 2021, 27(2): 592-603. DOI:10.1111/anu.13209
[17]
XU N, DING T, LIU Y T, et al. Effects of dietary tributyrin on growth performance, body composition, serum biochemical indexes and lipid metabolism-related genes expression of juvenile large yellow croaker (Larimichthys crocea) fed with high level soybean oil diets[J]. Aquaculture Nutrition, 2021, 27(2): 395-406. DOI:10.1111/anu.13192
[18]
XIE D Z, DAI Q Y, XU C, et al. Dietary tributyrin modifies intestinal function by altering morphology, gene expression and microbiota profile in common carp (Cyprinus carpio) fed all-plant diets[J]. Aquaculture Nutrition, 2021, 27(2): 439-453. DOI:10.1111/anu.13197
[19]
LEONEL A J, SILVA E L, AGUILAR E C, et al. Systemic administration of a nanoemulsion with tributyrin reduces inflammation in experimental colitis[J]. European Journal of Lipid Science and Technology, 2016, 118(2): 157-164. DOI:10.1002/ejlt.201400359
[20]
姜雪梅. 金霉素对猪肠道和粪便菌群的影响及抗性基因的迁移变化研究[D]. 硕士学位论文. 杭州: 浙江大学, 2020.
JIANG X M. Effects of chlortetracycline on the microbiota of intestine and feces in pigs and migration of antibiotic resistance genes[D]. Master's Thesis. Hangzhou: Zhejiang University, 2020. (in Chinese)
[21]
齐宇, 王开, 伊淑帅, 等. 产肠毒素大肠杆菌K88诱发BALB/c鼠肠炎模型的建立及评价[J]. 中国预防兽医学报, 2016, 38(1): 19-22.
QI Y, WANG K, YI S S, et al. Establishment and evaluation of bacterial enteritis model in BALB/c mice induced by enterotoxigenic Escherichia coli K88[J]. Chinese Journal of Preventive Veterinary Medicine, 2016, 38(1): 19-22 (in Chinese). DOI:10.3969/j.issn.1008-0589.2016.01.05
[22]
SWAGGERTY C L, PEVZNER I Y, KOGUT M H. Selection for pro-inflammatory mediators produces chickens more resistant to Eimeria tenella[J]. Poultry Science, 2015, 94(1): 37-42. DOI:10.3382/ps/peu053
[23]
朱荣生, 徐伟, 王怀中, 等. 饲粮添加不同水平三丁酸甘油酯对断奶仔猪生长性能、血清生化指标、肠组织形态和养分消化率的影响[J]. 动物营养学报, 2020, 32(2): 664-673.
ZHU R S, XU W, WANG H Z, et al. Effects of different supplemental levels of tributyrin on growth performance, serum biochemical indices, intestinal morphology and nutrient digestibility of weaned piglets[J]. Chinese Journal of Animal Nutrition, 2020, 32(2): 664-673 (in Chinese). DOI:10.3969/j.issn.1006-267x.2020.02.022
[24]
WANG C C, CAO S T, ZHANG Q H, et al. Dietary tributyrin attenuates intestinal inflammation, enhances mitochondrial function, and induces mitophagy in piglets challenged with diquat[J]. Journal of Agricultural and Food Chemistry, 2019, 67(5): 1409-1417. DOI:10.1021/acs.jafc.8b06208
[25]
邱洪仪. 三丁酸甘油酯和包被丁酸钠对慢性应激肉鸡肠道形态结构、二糖酶活性及脂质代谢的影响[D]. 硕士学位论文. 武汉: 武汉轻工大学, 2013.
QIU H Y. The effect of tributyrin and sodium butyrate on intestinal morphological structure, mucosa disaccharidase activity and lipid metabolism in broilers challenged with lipopolysaccharide[D]. Master's Thesis. Wuhan: Wuhan Polytechnic University, 2013. (in Chinese)
[26]
冯焱, 张芬鹊, 李建慧. 家禽肠道黏膜屏障结构及功能研究进展[J]. 中国家禽, 2016, 38(4): 1-4.
FENG Y, ZHANG F Q, LI J H. Research progress on the structure and function of poultry intestinal mucosal barrier[J]. China Poultry, 2016, 38(4): 1-4 (in Chinese).
[27]
肖武强, 徐敏丹, 吴先正. 脓毒症患者血清肠型脂肪酸结合蛋白、二胺氧化酶水平检测对早期肠组织损伤及预后的评估价值[J]. 现代检验医学杂志, 2021, 36(1): 10-13, 140.
XIAO W Q, XU M D, WU X Z. Evaluation value of serum intestinal fatty acid binding protein and diamine oxidase in the early stage of intestinal tissue injury and prognosis in patients with sepsis[J]. Journal of Modern Laboratory Medicine, 2021, 36(1): 10-13, 140 (in Chinese). DOI:10.3969/j.issn.1671-7414.2021.01.003
[28]
HODZIC Z, BOLOCK A M, GOOD M. The role of mucosal immunity in the pathogenesis of necrotizing enterocolitis[J]. Frontiers in Pediatrics, 2017, 5: 40.
[29]
CRIVELLI C, DEMARTA A, PEDUZZI R. Intestinal secretory immunoglobulin A (sIgA) response to Aeromonas exoproteins in patients with naturally acquired Aeromonas diarrhea[J]. FEMS Immunology and Medical Microbiology, 2001, 30(1): 31-35. DOI:10.1111/j.1574-695X.2001.tb01546.x
[30]
阎军红. 二胺氧化酶在小肠疾病诊断和治疗中的意义[J]. 国外医学(消化系疾病分册), 1994(4): 212-213.
YAN J H. The significance of diamine oxidase in the diagnosis and treatment of small intestinal diseases[J]. Foreign Medical Science (Digestive Diseases), 1994(4): 212-213 (in Chinese).
[31]
DAVIN-REGLI A, GUERIN-FAUBLÉE V, PAGÈS J M. Modification of outer membrane permeability and alteration of LPS in veterinary enterotoxigenic Escherichia coli[J]. Research in Veterinary Science, 2019, 124: 321-327. DOI:10.1016/j.rvsc.2019.04.011
[32]
吕勇, 尹峰, 薛月玲, 等. 内毒素、降钙素原及C反应蛋白检测在PICU细菌感染早期联合诊断的价值[Z]. 泰安: 泰安市妇幼保健院, 2016.
LV Y, YIN F, XUE Y L, et al. The value of detection of endotoxin, procalcitonin, and C-reactive protein in the early diagnosis of PICU bacterial infection[Z]. Taian: Tai'an Maternal and Child Health Hospital, 2016. (in Chinese)
[33]
宋建明. 评价血清降钙素原、C反应蛋白和血常规联合检测在重症细菌感染性疾病早期诊断中的应用价值[J]. 医学食疗与健康, 2020, 18(17): 182, 201.
SONG J M. To evaluate the application value of combined detection of serum procalcitonin, C-reactive protein, and routine blood in the early diagnosis of severe bacterial infectious diseases[J]. Medical Diet and Health, 2020, 18(17): 182, 201 (in Chinese).
[34]
熊珍, 张琼, 李樱. CRP、PCT诊断新生儿细菌感染性疾病的临床价值[J]. 中国继续医学教育, 2021, 13(6): 110-113.
XIONG Z, ZHANG Q, LI Y. Clinical value of CRP, PCT in the diagnosis of bacterial infectious diseases of newborn[J]. China Continuing Medical Education, 2021, 13(6): 110-113 (in Chinese). DOI:10.3969/j.issn.1674-9308.2021.06.031
[35]
关莹莹, 李云慧. 全血C反应蛋白联合血常规检验在细菌性感染性疾病诊断中的应用效果[J]. 中国实用医药, 2021, 16(8): 61-63.
GUAN Y Y, LI Y H. Application effect of whole blood C-reactive protein combined with blood routine test in the diagnosis of bacterial infectious diseases[J]. China Practical Medical, 2021, 16(8): 61-63 (in Chinese).