肥胖患者HFA小鼠模型的建立

目的研究肥胖患者的肠道菌群在无菌小鼠体内的定植规律。方法选取20只无菌KM小鼠,接种肥胖患者的粪便,构建菌群人源化(HFA)动物模型,利用变性梯度凝胶电泳技术(DGGE)评价患者肠道菌群在无菌小鼠体内的定植规律。结果 HFA小鼠菌群平均丰富度(richness,S)为12.04±3.68,肥胖患者的条带S为24,为HFA小鼠S的2倍;肥胖患者Shannon指数(H')为3.02,HFA小鼠平均H'为2.46±0.33;HFA小鼠与人肠道菌群的总相似度为26%;大部分雌性HFA小鼠与雄性HFA小鼠在聚类分析图上分离,且雄性HFA小鼠与患者更为相似。结论HFA小鼠体内能部分模拟肥胖患者的微生物区系,且与患者性别相同的小鼠模拟得更好。本实验建立的HFA模型为肥胖与肠道菌群关系的进一步研究提供新的选择。     

建立阿尔茨海默症人源肠道菌群动物模型

目的 使用粪菌移植方法 经口途径获得阿尔茨海默症人源肠道菌群(human flora-associated,HFA)模型小鼠,并通过生物信息学、组织病理学等方法对模型进行评价.方法 无菌雌性C57BL/6J小鼠28只,随机分为阿尔茨海默症模型(AD)组和对照(CON)组,分别经口途径接种阿尔茨海默症患者及健康志愿者的新...     

重症肌无力小鼠模型的建立

目的建立重症肌无力小鼠模型。方法电鳗乙酰胆碱受体(TnAChR)和完全弗氏佐剂(CFA)混合物免疫C57BL/6J小鼠,经二次加强免疫后,检测肌力、腓肠肌肌电图、膈肌终板电镜、血清抗体水平等指标。结果与对照小鼠相比,发病小鼠表现出肌力减弱的症状,肌电图显示小鼠腓肠肌复合动作电位振幅幅度显著下降,电镜证实神经肌接头处突触后膜变平、皱褶减少、空泡样变性,发病小鼠血清抗体水平明显升高。结论成功建立了重症肌无力小鼠模型,有助于探讨其发病机理及探索治疗自身免疫病新的途径和方法。     

阿莫西林干预IHFA小鼠及对其成年后肠道菌群的影响

目的利用婴儿菌群人源化小鼠(IHFA小鼠)观察阿莫西林对其干预后及成年后肠道菌群的影响。方法新生Balb/c无菌小鼠接种纯母乳喂养的婴儿粪便获得IHFA小鼠。7~21日龄灌胃给予100mg/kg阿莫西林,对照组给予等量的生理盐水。采用变性梯度凝胶电泳(DGGE)检测小鼠在21日龄及53日龄的肠道菌群。结果 21日龄的阿莫西林处理组IHFA小鼠肠道菌群与正常对照组比较差异有统计学意义(P<0.05);即使在停药后饲养至53日龄的成年小鼠,阿莫西林处理组小鼠肠道菌群仍然存在细微差异。结论哺乳期治疗剂量阿莫西林处理不仅严重干扰小鼠肠道菌群结构,同时导致其成年后肠道菌群不能完全恢复。     

APP/PS1转基因老年性痴呆模型小鼠肠道甲醛浓度异常升高

【目的】肠道菌群通过"微生物-肠道-脑轴"影响中枢神经系统的功能,同时也与老年性痴呆的发生发展相关,特别是盲肠内微生物菌群的变化更为显著。肠道菌群可以产生和代谢甲醛,而肠道能够迅速吸收甲醛;体内甲醛含量与老年性痴呆病人的认知损害程度呈正相关。因此,本文比较了7月龄APP/PS1转基因老年性痴呆模型小鼠(简称APP/PS1转基因小鼠)与同月龄C57BL/6J野生型小鼠(简称C57BL/6J小鼠)肠道菌群产生甲醛的情况。【方法】取APP/PS1转基因小鼠(n=8)与C57BL/6J小鼠(n=9)的不同肠段(十二指肠、小肠、盲肠、结肠),采用2,4-Dinitrophenylhydrazi zne(DNPH)显色偶联高效液相色谱(HPLC coupled with DNPH)测定肠道消化物和肠壁组织的甲醛。【结果】APP/PS1转基因小鼠盲肠消化物内的甲醛含量,较C57BL/6J小鼠存在显著升高(P=0.036);而两者小肠和结肠消化物甲醛含量无显著差别。在两种小鼠之间,小肠壁内甲醛存在差异(P=0.052),而盲肠和结肠壁甲醛含量无显著差异(P0.05)。【结论】肠道菌群是小鼠体内甲醛的主要来源之一,无论肠道消化物,还是肠道壁组织均为盲肠的甲醛含量最高。这些结果表明,APP/PS1转基因小鼠肠道菌群存在甲醛代谢失调,从而导致其肠道消化物的甲醛含量升高。     

水溶性壳聚糖对小鼠肠道菌群的影响

目的 应用聚合酶链式反应-变性梯度凝胶电泳（PCR-DGGE）技术研究水溶性壳聚糖对正常小鼠肠道菌群的影响。方法 使用SPF级昆明小鼠18只,随机分为3组,每组6只,依次为正常对照组（NC）、水溶性壳聚糖高（WSC-H）和低浓度组（WSC-L）,灌服对应药物30 d后收取鼠便,细菌基因组DNA提取,PCR-DGGE电泳得到肠道菌群图谱,计算菌群结构多样性指数,聚类分析相似性并鉴定优势条带序列。结果 水溶性壳聚糖处理后小鼠肠道菌群结构发生改变,多样性指数、丰富度和均匀度较低,乳杆菌成为肠道中的优势菌群。结论 水溶性壳聚糖能够增加肠道中乳杆菌的种类和数量,起到益生元的作用来调节肠道微生态平衡。     

C57BL／6J小鼠ES细胞系的建立及其特性分析

本文报道从C57BL/6J个鼠的囊胚中,建立了三个ES细胞系MESPU17,MESPU18,MESPU19。这些细胞的细胞学特征和强AKP反应,表明这三个细胞系具有干细胞的特征。这三个细胞系均为XY型,正常二倍体核型分别占70％、88％和59％。体外分化可形成简单类胚,体内分化可形成瘤块。与国际上通用的CCE和来自129/ter的ES细胞MESPU13相比,这三个细胞系的ES细胞较大;在体外培养时,生长较慢;细胞也较粘,进行显微注射时,很容易粘在注射针壁上。MESPU17,MESPU18进行了嵌合体制作,以BALB/c和昆明鼠的囊胚为受体,采用囊胚注射法未获嵌合体,但使用昆阴鼠的8细胞胚注射法和共培养法得到嵌合体。     

肠道菌群变化对实验小鼠肠黏膜免疫的影响

目的探讨肠道菌群变化对肠黏膜相关淋巴组织的影响。方法通过变性梯度凝胶电泳（Denatu-ring gradient gel electrophoresis,DGGE）法研究了三种不同级别实验小鼠即清洁级小鼠、SPF小鼠和普通小鼠肠道菌群的组成,并用免疫组织化学（immunohistochemistry,IHC）方法研究了此三种不同级别的实验小鼠肠黏膜相关淋巴组织sIgA阳性细胞分布情况。结果普通小鼠肠道细菌种类最多,其sIgA阳性细胞分布最多,肠道不同部位之间sIgA分布情况差异有显著性（P〈0.05）,小肠和大肠之间的阳性细胞分布差异极显著（P〈0.01）;其次是清洁级小鼠,其肠道不同部位之间菌种组成差异无显著性,小肠和大肠之间的阳性细胞分布差异有显著性（P〈0.05）;SPF小鼠肠道细菌种类最少,故其sIgA阳性细胞分布最少,且其肠道不同部位之间菌种组成差异无显著性,小肠和大肠之间的阳性细胞分布差异无显著性（P〉0.05）。结论随着动物微生物控制级别的增高,肠道微生物多样性递减;sIgA阳性细胞与肠道细菌种类正相关。     

螺旋藻对腹泻模型小鼠肠道菌群的影响

为评价螺旋藻对肠道菌群的影响, 采用注射氨苄青霉素的方法, 制造小鼠腹泻模型, 造模成功后灌服不同剂量螺旋藻进行治疗, 取不同治疗时间的粪便样品, 检测双歧杆菌、乳杆菌、大肠杆菌、肠球菌、拟杆菌、产气荚膜梭菌菌群的数量,并在螺旋藻灌胃8 d后, 测定小鼠不同肠段各指标菌群的数量。结果表明, 中高剂量组螺旋藻对腹泻模型小鼠肠道菌群有明显调整作用, 有效缩短了肠道菌群由失调到平衡的时间;粪便样品同各肠段内容物所反映的菌群的变化趋势相同, 即治疗恢复组的双歧杆菌、乳杆菌数量总体上高于生理盐水组, 肠杆菌、肠球菌、拟杆菌、产气荚膜梭菌数量均低于生理盐水组, 但取样位置不同, 各指标菌群的数量明显存在差异。     

转Bar基因稻谷对小鼠(Mus musculus)肠道菌群的作用研究

选取6周龄、雌雄各半、体重18~22 g的SPF级昆明小鼠(Mus musculus)100只,随机分成5组,每组4个重复,每个重复5只小鼠。分别用含量为40%和60%的转Bar基因稻谷Bar68-1和非转基因稻谷D68日粮喂养小鼠,亲代小鼠饲养180 d后开始繁殖子一代(F1),每代小鼠饲养180 d。在180 d后分别从亲代和子一代每组随机抽取5只小鼠,提取肠道内容物基因组DNA,利用细菌通用引物对细菌16S r DNA的V3区序列进行PCR扩增,并将扩增产物用变性梯度凝胶电泳(denaturing gradient gel electrophoresis DGGE)。研究显示,各组小鼠肠道菌群相似性系数无显著差异(P0.05);肠道主要优势菌群相同。结果表明,转Bar基因稻谷对小鼠肠道菌群的作用不明显。     

Establishment and development of intestinal microbiota in preterm neonates

Microbial colonization of the infant gut is essential for the development of the intestine and the immune system. The profile of intestinal microbiota in the full-term, vaginally delivered, breast-fed infant is considered as ideally healthy. However, in preterm infants this process is challenging, mainly because of organ immaturity, antibiotics use, and hospital stay. To assist in a proper microbiota development in these infants, a detailed knowledge of the colonization process, and the differences from that of full-term breast-fed infants, is needed. We assessed the establishment of the gut microbiota and its metabolic activity in preterm neonates (n?=?21) during the first 3?months of life and compared it with that of vaginally delivered, exclusively breast-fed full-term infants (n?=?20) using qualitative and quantitative culture-independent methods. Differences in the gut microbiota composition between both groups were observed. Preterm infants showed higher levels of facultative anaerobic microorganisms and reduced levels of strict anaerobes such as Bifidobacterium, Bacteroides, and Atopobium. Short-chain fatty acids concentrations were lower in preterm infants during the first days of life. Alterations occur in the process of microbiota establishment in preterm infants, indicating the need for intervention strategies to counteract them.     

Lactobacillus strains stabilize intestinal microbiota in Japanese cedar pollinosis patients

A randomized double-blind, placebo-controlled trial was conducted to ascertain the intestinal microbiota-altering properties of LGG and L. gasseri TMC0356 (TMC0356) in Japanese cedar Cryptomeria japonica pollinosis patients. Fecal bacteria communities were examined before and after fermented milk administration using culture, FISH and T-RFLP methods. Test group subjects showed the presence of LGG and TMC0356 along with a significant increase in fecal lactobacilli ( P < 0.001) after giving LGG and TMC0356 fermented milk. Culture and FISH analysis revealed no significant changes in other intestinal bacterial groups. Each subject exhibited a characteristic T-RFLP profile pattern that varied quantitatively and qualitatively with JCP shedding. Profile changes were observed in 53% of placebo group subjects and in 21% of test group subject's post-administration, indicating that LGG and TMC0356 suppressed intestinal microbiota changes in JCPsis patients. The results suggest that intestinal microbiota might be more sensitive to exposure to environmental allergens than expected from the results of general culture method studies. Stabilization of intestinal microbiota by selected probiotic strains such as LGG and TMC0356 could be beneficial to homeostasis of the intestinal microbiota and useful in the management of JCPsis.     

Establishment of inbred strains of chicken and Japanese quail and their potential as animal models.

We started establishing inbred strains of chicken and Japanese quail in 1970. In class Aves, full sib mating is highly difficult due to inbreeding depression. In the chicken, we attempted to establish some inbred strains in three breeds, Black Minorca, White Leghorn and Fayoumi by fixing all the characters that differentiate individuals homozygously. In this paper, we describe some marker genes and characters fixed in the inbred strains of chicken and Japanese quail as well as a calculation of a putative coefficient of inbreeding in 8 chicken inbred strains using band sharing values detected by AFLP analysis. We established generalized glycogenosis type II quail, myotonic dystrophy quail, neurofilament deficient quail, visually impaired chicken, double oviduct chicken with partial kidney deficiency, chicken showing spontaneous lymphocytic thyroiditis with feathered amelanosis, and chicken with a hereditary nervous disorder. The processes of establishment and characteristics of these animal models are described with some interesting information obtained from these animal models. In generalized glycogenosis type II quail, the results of enzyme replacement therapy and gene therapy are described.     

口服肝素与小鼠肠道菌群的相互作用

口服肝素药物的开发需要系统地理解口服肝素与肠道菌群之间的互作过程。通过荧光体视镜观察荧光素标记的肝素经小鼠口服后在体内的分布情况,利用高效液相色谱法检测肝素在模拟胃肠液中的稳定性和体外培养肠道菌群模拟肠道菌对肝素的降解作用,发现口服肝素主要分布在小鼠胃肠道内,在体外模拟胃肠液条件下肝素结构稳定,但能够被添加肝素的厌氧培养基培养后的肠道菌群降解。为了进一步揭示口服肝素对健康小鼠肠道菌群的影响,利用Illumina MiSeq高通量测序技术测定口服肝素后C57BL/6J小鼠粪便菌群的16S rRNA序列,与口服生理盐水的小鼠粪便菌群进行对比,发现口服肝素的小鼠粪便菌群的生物多样性降低;在门水平上,菌群结构差异不显著;而在属水平上,别样杆菌属Alistipes、副萨特氏菌属Parasutterella和艾克曼菌属Akkermansia相对丰度增高,而嗜胆菌属Bilophila、肠杆菌属Enterorhabdus、瘤胃梭菌属Ruminiclostridium、普雷沃氏菌科Prevotellaceae_UCG_001、瘤胃梭菌属Ruminiclostridium-9、拟杆菌属Bacteroides、Lachnoclostridium、Candidatus_Saccharimonas、Intestinimonas和Dubosiella的相对丰度减少,表明口服肝素能够影响小鼠肠道菌群结构。此外,实验发现口服肝素对小鼠无明显毒副作用,具有较高安全性。研究结果将为开发肝素口服递送策略提供新的思路,为口服肝素类药物的开发提供参考。     

鸡肠道微生物菌群的建立发育、分布和生理学意义

鸡的胃肠道具有复杂的微生物菌群,该微生物菌群与宿主的肠道和整体健康密切相关,为了全面揭示鸡肠道微生物菌群的组成及其功能,本文对鸡肠道微生物菌群的建立发育、各肠段群落的分布及其生理学意义进行综述,从而为鸡肠道功能菌株的分离及有效利用,合理调控微生物菌群-宿主相互作用,提高饲料转化率和改善肠道健康提供理论依据。     

Leeches and their microbiota: naturally simple symbiosis models

Strictly blood-feeding leeches and their limited microbiota provide natural and powerful model systems to examine symbiosis. Blood is devoid of essential nutrients and it is thought that symbiotic bacteria synthesize these for the host. In this review, three distinct leech-microbe associations are described: (i) the mycetome, which is the large symbiont-containing organ associated with the esophagus; (ii) the nephridia and bladders that form the excretory system; and (iii) the digestive tract, where two bacterial species dominate the microbiota. The current knowledge and features of leech biology that promote the investigation of interspecific interactions (host-microbe and microbe-microbe) and their evolution are highlighted.     

Screening of exopolysaccharide-producing Lactobacillus and Bifidobacterium strains isolated from the human intestinal microbiota

Using phenotypic approaches, we have detected that 17% of human intestinal Lactobacillus and Bifidobacterium strains could be exopolysaccharide (EPS) producers. However, PCR techniques showed that only 7% harbored genes related to the synthesis of heteropolysaccharides. This is the first work to screen the human intestinal ecosystem for the detection of EPS-producing strains.     

Long-term evolution and short-term adaptation of microbiota strains and sub-strains in mice

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A comparison of their virulence for mice and the enzymatic activity of Streptococcus pneumoniae strains

In most cases no correlation between the virulence of S. pneumoniae and their enzymatic activity was registered in 101 S. pneumoniae strains isolated in pneumococcal infections of different localization. Pneumococcal strains belonging to different serotypes and characterized by their low virulence for mice (LD50 = 10(6) colony-forming units) had the highest neuraminidase and protease-alzolase activity in comparison with highly virulent cultures of these bacteria. In pneumococcal cultures in the R-form avirulence for mice occurred mainly in combination with low enzymatic activity.