摘要
目的探究Akkermansia muciniphila(AKK)对葡聚糖硫酸钠溶液(DSS)诱导的小鼠模型的影响。方法无特定病原体(SPF)级C57BL/C雄性小鼠30只,依次分为空白组、结肠炎组、AKK+结肠炎组。观察每组小鼠的日常活动量、排便情况,选择疾病活动指数(DAI)进行疾病活动度评估,收集粪便行粪便做隐血试验等,在建模结束7 d后采用断头法处死小鼠,酶联免疫吸附试验(ELISA)检测结肠组织白细胞介素(IL)-1β、IL-6、肿瘤坏死因子-α(TNF-α)含量,苏木精-伊红(HE)检测结肠病理结构,蛋白质印迹法(Western blot)检测结肠黏膜中紧密连接蛋白Occludin-1的表达,二喹啉甲酸(BCA)法测定结肠组织二胺氧化酶(DAO)、D-乳酸的浓度。采用单因素方差分析(one-way ANOVA)进行统计分析,组间两两比较用LSD-t或Tamhane’s T2法进行显著性分析。结果第3、7、21天空白组的体重分别为(17.01±1.30)、(19.77±1.85)、(21.07±1.80)g,结肠炎组的体重分别为(15.67±0.65)、(13.66±0.62)、(11.72±0.53)g,AKK+结肠炎组的体重分别为(15.74±1.43)、(15.14±1.31)、(13.37±1.46)g,第3天结肠炎组的体重有低于AKK+结肠炎组的趋势,但差异无统计学意义(LSD-t=0.133,P>0.05),第7天及第21天时结肠炎组及AKK+结肠炎组体重显著低于空白对照组(LSD-t=10.061、15.272,P<0.01),结肠炎组低于AKK+结肠炎组(LSD-t=2.436、2.706,P<0.05)。第3、7、21天空白组的DAI指数均为0,结肠炎组的DAI指数分别为1.25±0.06、2.26±0.13、3.13±0.08,AKK+结肠炎组的DAI指数分别为0.89±0.43、1.53±0.05、2.45±0.10,AKK+结肠炎组DAI评分低于结肠炎组(LSD-t=3.242、20.444、2.027,P<0.05)。HE结果显示结肠炎组隐窝排列不规则、炎症细胞浸润、肠腺水肿,AKK菌灌胃后,结肠水肿和出血减轻,肠腺结构清晰,空白组、结肠炎组、AKK+结肠炎组的病理评分分别为(0.76±0.03)、(4.07±0.05)、(2.82±1.03)分,AKK+结肠炎组高于空白组、低于结肠炎组(LSD-t=13.208、5.985,P<0.05)。结肠炎组中IL-1β、IL-6、TNF-α分别是(8.61±3.06)、(5.58±2.23)、(55.68±18.57)pg/ml,明显高于空白组(Tamhane’s T2=6.584、7.175、9.239,P<0.01),与结肠炎组相比,AKK+结肠炎组IL-1β、IL-6、TNF-α表达下降,分别是(5.51±2.32)、(3.12±0.89)、(20.94±8.05)pg/ml(Tamhane’s T2=3.047、3.667、6.617,P<0.05)。结肠炎组中Occludin-1、D-乳酸、DAO分别是(615.56±41.87)ng/ml、(4.37±0.15)μg/ml、(8.93±0.07)ng/ml,明显高于空白组(Tamhane’s T2=19.059、36.071、23.676,P<0.01),与结肠炎组比较,AKK+结肠炎组Occludin-1、D-乳酸、DAO分别是(887.55±42.48)ng/ml、(3.57±0.20)μg/ml、(7.41±0.07)ng/ml。与结肠炎组相比,差异有统计学意义(Tamhane’s T2=14.421、47.183、1.020,P<0.01)。结论AKK菌可降低IL-1β、IL-6、TNF-α含量,通过调控Occludin-1等结肠紧密连接蛋白,从而降低结肠炎的严重程度,增强结肠的屏障功能。
Objective To investigate the effects of Akkermansia muciniphila(AKK)on mouse models induced by dextran sulfate sodium(DSS).Methods Totally,30 specific pathogen free(SPF)C57BL/C male mice were divided into blank group,colitis group and AKK+colitis group.Daily activity and defecation of mice in each group were observed.Disease active index(DAI)was selected for disease activity assessment,feces were collected for occult blood test.At 7th day,mice were killed by decapitation method.The contents of interleukin(IL)-1β,IL-6 and tumor necrosis factor-α(TNF-α)in colon tissue were determined by enzyme linked immunosorbent assay(ELISA).The pathological structure of colon was deter-mined by hematoxylin and eosin(HE)staing.The expression of Occludin-1 in colon mucosa was detected by Western blotting.The concentration of diamine oxidase(DAO)and D-lactic acid in colon tissue was deter-mined by bicinchoninic acid(BCA).One-way ANOVA was used for statistical analysis,and LSD-t or Tam-hane's T2 method was used for pairwise comparison between groups for significance analysis.Results The body mass in the blank group at the 3rd,7th and 21st day was(17.01±1.30),(19.77±1.85)and(21.07±1.80)g,that in the colitis group was(15.67±0.65),(13.66±0.62)and(11.72±0.53)g,and that in AKK+colitis group was(15.74±1.43),(15.14±1.31)and(13.37±1.46)g.The body mass in AKK+colitis group was lower than that in colitis group at the 3rd day,but the difference was not statistically significant(LSD-t=0.133,P>0.05).Body mass in the colitis group and AKK+colitis group at the 7th and 21st day was significantly lower than that in the blank control group(LSD-t=10.061,15.272,P<0.001),colitis group was lower than AKK+colitis group(LSD-t=2.436,2.706,P<0.05).The DAI index of the blank group at the 3rd,7th and 21st day was O,that of the colitis group was 1.25±0.06,2.26±0.13 and 3.13±0.08,and that of AKK+colitis group was 0.89±0.43,1.53±0.05 and 2.45±0.10,respectively.The DAI in AKK+colitis group was higher than in blank group and lower than in colitis group(LSD-t=3.242,20.444,2.027,P<0.05).HE staining showed that the coli-tis group had irregular crypt arrangement,inflammatory cell infiltration and intestinal glandular edema.After intragastric administration of AKK,colonic edema and bleeding were alleviated,and intestinal glan-dular structure was clear.The pathological scores in blank group,colitis group and AKK+colitis group were(0.76±0.03),(4.07±0.05)and(2.82±1.03)respectively.The pathological scores in AKK+colitis group were higher than in blank group and lower than in colitis group(LSD-t=13.208,5.985,P<0.05).The levels of IL-1β,IL-6 and TNF-αin the colitis group were(8.61±3.06),(5.58±2.23)and(55.68±18.57)pg/ml respectively,which were significantly higher than those in the blank group(Tamhane's T2=6.584,7.175,9.239,P<0.001).Compared with the colitis group,the expression levels of IL-1β,IL-6 and TNF-αin the AKK+colitis group were decreased(5.51±2.32),(3.12±0.89),(20.94±8.05)pg/ml,Tamhane's (T2=3.047,3.667,6.617,P<0.05).0ccludin-1,D-lac-tic acid and DA0 in colitis group were(615.56±41.87)ng/ml,(4.37±0.15)μg/ml and(8.93±0.07)ng/ml,significantly higher than those in blank group(Tamhane's T2=19.059,36.071,23.676,P<0.001).Compared with colitis group,Occludin-1,D-lactic acid and DAO in AKK+colitis group were(887.55±42.48)ng/ml,(3.57±0.20)μg/ml and(7.41±0.07)ng/ml,respectively.Compared with colitis group,there was a statistically significant difference(Tamhane's T2=14.421,47.183,1.020,P<0.001).Conclusion AKK can decrease the contents of IL-1β,IL-6 and TNF-α,and regu-late Occludin-1 and other colonic tight junction proteins,thereby reducing the severity of colitis and enhan-cing the barrier function of colon.
作者
张琳
魏凯艳
胡惠祥
郑建涛
刘益娟
Zhang Lin;WWei Kaiyan;Hu Huixiagng;Zheng Jiantao;LiuYjuan(Department of Gastrointestinal Surgery,National Regional Medical Center,Binhai Campus of the First Afiliated Hospital of Fujian Medical University,Fuzhou 350212,China;Department of Gastrointestinal Surgery,the First Affliated Hospital of Fujian Medical University,Fuzhou 350005,China;The First Clini cal Medical College of Fujan Medical University,Fuzhou 350005,China;Department of Gastroenterology,National Regional Medical Center,Binhai Campus of the First Affiliated Hospital of Fujian Medical University,Fuzhou 350212,China;Department of Gastroenterology,the First Affiliated Hospital of Fujian MedicalUniversity,Fuzhou350005,China)
出处
《中华实验外科杂志》
CAS
北大核心
2023年第5期867-872,共6页
Chinese Journal of Experimental Surgery
基金
福建省科技厅创双高联合基金(2019Y9120)
福建省自然科学基金(2020J01977)。
关键词
结肠炎
炎性因子
结肠紧密连接蛋白
黏膜屏障
Colitis
Inflammatory cytokines
Colonic tight junction protein
Mucosal barrier
作者简介
通信作者:刘益娟,Email:liuyijuan0611@163.com。
