肝细胞FXR对日本血吸虫感染小鼠肠道菌群的影响

目的分析肝细胞法尼醇X受体(farnesoid X receptor,FXR)特异性敲除对日本血吸虫感染小鼠肠道菌群的影响。方法6~8周龄野生型雄性C57BL/6J和肝细胞FXR特异性敲除小鼠(FXR-HKO)随机分为4组:野生型小鼠正常对照组(WT,n=5)、FXR-HKO小鼠正常对照组(FXR-HKO,n=6)、野生型小鼠日本血吸虫感染组(Sj-WT,n=6)、FXR-HKO小鼠日本血吸虫感染组(Sj-FXR-HKO,n=5)。其中,感染组每只小鼠感染(15±1)条日本血吸虫尾蚴,感染第5周处死小鼠,在无菌条件下收取小鼠结肠内容物,进行16S rDNA测序,比较各组之间的肠道菌群多样性和丰度差异。结果Beta多样性分析结果显示,Sj-WT组和Sj-FXR-HKO组小鼠的肠道菌群与正常小鼠肠道菌群有明显的分群。在门水平上,与正常对照组相比,Sj-WT组和Sj-FXR-HKO组小鼠肠道中拟杆菌门的丰度升高,厚壁菌门丰度降低,厚壁菌门与拟杆菌门丰度比值明显降低(均P<0.05)。在属水平上,与正常对照组相比,日本血吸虫感染后小鼠肠道中拟杆菌属的丰度明显升高(均P<0.05),Sj-FXR-HKO组拟杆菌属的丰度比Sj-WT组的升高更明显(P<0.05),其中Sj-WT组脱硫弧菌属、杜氏菌属、乳杆菌属的丰度显著降低(均P<0.05),Sj-FXR-HKO组瘤胃球菌科属、毛罗菌科、杜氏菌属、乳杆菌属的丰度明显降低(均P<0.05)。结论肝细胞FXR特异性敲除影响了日本血吸虫感染小鼠肠道菌群稳态,为后续进一步研究FXR-肠道菌群在血吸虫病中的作用及机制奠定了扎实的实验基础。

一种裂体吸虫与日本血吸虫蛋白质差异研究

目的 探讨裂体吸虫 x(S.x)和日本血吸虫 (S.j)的蛋白质有无差异。方法 采用 SDS-PAGE法。感染了 S.x和 S.j尾蚴的兔 45 d后剖杀 ,从肠系膜静脉血管中取出成虫 ,各 2 0对 ,匀浆 ,高速离心后取上清液 ,电泳 ,分离胶和浓缩胶分别为 10 %和 4%。电泳完成后以考马斯兰 R2 50 染色。应用分子分析软件系统 Gel Doc10 0 0 (BIO-RAD)进行分析 ,并计算出 S.x与 S.j的比值 (S.x/ S.j)。结果 S.x和 S.j蛋白带各有 2 6条 ,其中 3 5 .0 k Da和 10 9.8k Da为 S.x和 S.j分别特有的蛋白带 ,它们之间存在明显的差异 ;S.x和 S.j的 87.3 k Da,40 .9k Da和 3 2 .2 k Da蛋白质量差异达 2倍。 15 .2 k Da蛋白质量差异 1.9倍 ,73 .2 k Da蛋白质量差异 1.7倍。结论 S.x和 S.j成虫可溶性蛋白所分出的带数经定性、定量分析 。

AUTORADIOGRAPHIC STUDY OF THE ATTRITION OF MIGRATING SCHISTOSOMULA IN THE SKIN OF MICE II.MICE CHALLENGED WITH NORMAL S.MANSONI CERCARIAE AFTER IMMUNIZATION WITH HIGHLY IRRADIATED CERCARIAE

The migratory pattern and attrition of75Se-labelled S.mansoni challenge cercariae in CF1mice immunized 0,1,3, or 5 times with 12kR-or 48kR-irradiated cercariae were investigatedby compressed organ autoradiography.Mice were percutaneously inoculated with 500-1000 cer-cariae for each immunization and with a known count of approximately 90 cercariae at adifferent site for challenge.The skin,lungs,liver,other organs,and catrcass were processedfor autoradiography.Hepatic and mesenteric perfusions for worm collection were alsosampled from day 19 through 34.In naive tnice,25% of challenge cercariae died in theskin.Skin attrition of challenge cercariae in mice immunized with 12kR-irradiated cercariaewas 43% with 1 immunization,54% with 3,and 58% with 5,whereas in mice immunized with48kR-irradiated cercariae,skin,attrition was48% with 1,63% with 3,and 76% with 5 immunni-zations.In all immunized mice,a high percentage of migrating sehistosomula died in the skinwithin 24 hours of a challenge infection.Utilizing autoradiography,it was found that thesilver foci created by single labelled worms were difficult to differentiate from those of pairedworms.This study documents the induction of acquired resistance against a challenge infectionin the skin by vaccination with highly irradiated cercariae,with the degree of attemtation ofimmunizing cercariae and the number of immunizations being closely related to the attritiotn.rate in the skin.Finally,we discuss the contradictory views concerning the site of attritiotn ofchallenge cercariae.

日本血吸虫视黄酸X受体2全长cDNA的克隆及初步分析

目的克隆编码日本血吸虫视黄酸X受体2(SjRXR2)蛋白的全长cDNA,并对其进行初步研究。方法利用cDNA末端快速扩增技术(RACE)获得SjRXR2蛋白全长编码cDNA。利用生物信息学技术,对基因结构进行初步分析。利用实时荧光定量(Real time)PCR技术对该基因在日本血吸虫不同时期虫体中的转录情况进行分析。应用在线抗体表位预测软件获得SjRXR2配体结合区抗原性较强的一个多肽序列,合成该多肽片段,并免疫小鼠制备抗血清。利用Western blot技术分析该蛋白在日本血吸虫中的表达。结果采用RACE技术成功获得了SjRXR2蛋白全长编码cDNA,总长度为5 960bp,其完整开放阅读框为4 308 bp,编码1 435个氨基酸,预测分子量为159 kDa。生物信息学分析表明该基因编码的蛋白质序列具有核受体家族2的典型结构域特征,且与曼氏血吸虫RXR2有较高的相似性。Real time PCR分析表明,该基因在21、42 d龄日本血吸虫虫体内有较高的转录水平。Western blot分析表明,小鼠SjRXR2多肽免疫血清可特异性识别日本血吸虫虫体150 kDa蛋白。结论成功获得了编码SjRXR2蛋白的全长cDNA,并制备了针对该蛋白的特异性多克隆抗体,为进一步研究该蛋白的功能奠定了基础。