基于HMGB1-TLR2/TLR4-NF-κB通路探索氧化苦参碱对小鼠微小隐孢子虫感染的影响  

Effect of oxymatrine on Cryptosporidium parvum infection in mice based on the HMGB1-TLR2/TLR4-NF-κB pathway

作　　者：史洁 汲蕊 管志玉 张晓宁 逯一龙 SHI Jie;JI Rui;GUAN Zhiyu;ZHANG Xiaoning;LU Yilong(School of Basic Medical Sciences,Shandong Second Medical University,Weifang,Shandong 261053,China;College of Traditional Chinese Medicine,Shandong Second Medical University,Weifang,Shandong 261053,China)

机构地区：[1]山东第二医科大学基础医学院,山东潍坊261053 [2]山东省第二医科大学中医学院,山东潍坊261053

出　　处：《中国血吸虫病防治杂志》2024年第3期286-293,共8页Chinese Journal of Schistosomiasis Control

基　　金：山东省自然科学基金(ZR2020MH382);山东省自然科学基金青年项目(ZR2015HQ030)。

摘　　要：目的 探讨高迁移率族蛋白B1(high mobility group box protein B1,HMGB1)-Toll样受体2(Toll-like receptor 2,TLR2)/TLR4-核因子κB(nuclear factorκB,NF-κB)通路在微小隐孢子虫感染致肠黏膜损伤中的作用,以及氧化苦参碱(oxymatrine,OMT)对小鼠微小隐孢子虫感染的干预作用。方法 4周龄SPF级BALB/c小鼠40只随机分成4组,分别为对照组、感染组、甘草酸(glycyrrhizin,GA)组及OMT组。感染组、GA组及OMT组小鼠给予地塞米松免疫抑制1周后,每只灌胃1×10^(5)个微小隐孢子虫卵囊建立微小隐孢子虫肠道感染小鼠模型。模型建立成功后,GA组小鼠连续2周腹腔注射GA 25.9 mL/(kg·d),OMT组连续2周经口灌胃OMT 50 mg/(kg·d);对照组正常饮食、饮水。治疗2周后剖杀各组小鼠,取空肠近端组织。采用苏木精-伊红(hematoxylin-eosin,HE)染色观察小鼠肠黏膜病理变化,测量肠绒毛高度、肠隐窝深度及两者比值;采用免疫组织化学染色检测小鼠肠上皮细胞中闭合蛋白(occludin)和紧密粘连蛋白1(zonula occludens protein 1,ZO1)表达水平,采用实时荧光定量PCR(quantitative real-time PCR,qPCR)检测小鼠空肠组织中HMGB1、TLR2、TLR4、髓样分化因子88(myeloid differentiation primary response gene 88,MyD88)、NF-κB p65 mRNA相对表达量。结果 HE染色结果显示,与对照组比较,感染组小鼠肠绒毛明显萎缩变短、脱落,黏膜下层水肿;GA组和OMT组小鼠肠绒毛结构趋于完整,排列趋于整齐。各组小鼠肠绒毛高度(F=6.207,P=0.000 5)、肠隐窝深度(F=6.903,P=0.000 3)及两者比值(F=37.190,P <0.000 1)差异均有统计学意义。感染组小鼠肠绒毛高度[(321.9±41.1)μm]显著低于对照组[(399.5±30.9)μm](t=4.178,P <0.01)和GA组[(383.7±42.7)μm](t=3.130,P <0.01),感染组小鼠肠隐窝深度[(185.0±35.9)μm]显著高于对照组[(128.4±23.6)μm](t=3.877,P <0.01)及GA组[(143.3±24.7)μm](t=2.710,P <0.05)。OMT组小鼠肠绒毛高度[(375.3±22.9)μm]显著高于感染组(t=3.888,P <0.01)Objective To investigate the involvement of the high mobility group box protein B1 (HMGB1)-Toll-like receptor 2(TLR2)/TLR4-nuclear factorκB (NF-κB) pathway in the intestinal mucosal injury induced by Cryptosporidium parvum infection,and to examine the effect of oxymatrine (OMT) on C.parvum infection in mice.Methods Forty SPF 4-week-old BALB/c mice were randomly divided into four groups,including the control group,infection group,glycyrrhizin (GA) group and OMT group.Each mouse was orally administered with 1×10~5C.parvum oocysts one week in the infection,GA and OMT groups following dexamethasone-induced immunosuppression to model C.parvum intestinal infections in mice.Upon successful modeling,mice in the GA group were intraperitoneally injected with GA at a daily dose of 25.9 mL/kg for successive two weeks,and animals in the OMT group were orally administered OMT at a daily dose of 50 mg/kg for successive two weeks,while mice in the control group were given normal food and water.All mice were sacrificed two weeks post-treatment,and proximal jejunal tissues were sampled.The pathological changes of mouse intestinal mucosal specimens were observed using hematoxylin-eosin (HE) staining,and the mouse intestinal villous height,intestinal crypt depth and the ratio of intestinal villous height to intestinal crypt depth were measured.The occludin and zonula occludens protein 1 (ZO1) expression was determined in mouse intestinal epithelial cells using immunohistochemistry,and the relative expression of HMGB1,TLR2,TLR4,myeloid differentiation primary response gene 88(MyD88) and NF-κB p65 mRNA was quantified in mouse jejunal tissues using quantitative real-time PCR (qPCR) assay.Results HE staining showed that the mouse intestinal villi were obviously atrophic,shortened,and detached,and the submucosal layer of the mouse intestine was edematous in the infection group as compared with the control group,while the mouse intestinal villi tended to be structurally intact and neatly arranged in the GA and OMT groups.There were si

关 键 词：微小隐孢子虫 高迁移率族蛋白B1 Toll样受体 核因子κB 氧化苦参碱 甘草酸 肠黏膜屏障 小鼠 

分 类 号：R382.3[医药卫生—医学寄生虫学]