牦牛肠源Lactobacillus acidophilus L3对其小肠黏膜形态结构的影响

为了探讨牦牛肠源嗜酸乳杆菌Lactobacillus acidophilus L3对牦牛小肠肠道黏膜形态结构的影响,将10头健康牦牛(2~2.5岁)随机分为2组,分别为益生菌组和空白对照组,益生菌组在饲料中添加2×109CFU·kg-1L.acidophilus L3,饲喂28 d后取其小肠组织样品,采用组织学和显微测量学方法对益生菌组和空白对照组牦牛小肠绒毛高度、隐窝深度、绒毛高度/隐窝深度比值(V/C),以及回肠黏膜淋巴集结厚度进行测量和统计分析。研究结果显示,益生菌组牦牛十二指肠、空肠、回肠绒毛高度均较空白对照组显著增加,隐窝深度降低,V/C值增加,差异有统计学意义(P<0.05)。通过对回肠淋巴集结的测量发现,益生菌组牦牛回肠淋巴集结的平均厚度均较空白对照组显著增加,差异有统计学意义(P<0.05)。研究结果表明,牦牛肠源L.acidophilus L3可有效改善其肠道黏膜结构,增强肠道黏膜免疫效应部位的免疫功能。     

牦牛骨胶对小鼠应激及免疫机能的功能影响

目的：通过实验研究牦牛骨胶对小鼠应激及免疫功能的影响。方法：采用昆明小鼠为实验对象,观察小鼠负重游泳、耐缺氧、高温和低温存活时间,以及正常小鼠网状内皮系统吞噬能力和溶血素抗体水平。结果：灌胃牦牛骨胶能明显增强小鼠耐缺氧、耐寒、抗疲劳的能力,也能增强小鼠网状内皮系统吞噬功能、明显升高免疫受抑小鼠的HC50,具有显著的抗应激和增强免疫功能的作用。     

一株猪肠道Lactobacillus animalis LGM对结肠炎小鼠Th细胞分化转录因子基因表达的影响

【目的】从体外和体内研究Lactobacillus animalis LGM对Th细胞分化转录因子T-bet、GATA-3、ROR-γt和Foxp3的调节作用,以及探究L. animalis LGM对小鼠结肠炎的影响。【方法】本试验采用改良型的Hungate滚管技术从猪结肠内容物中分离一株L. animalis LGM,根据其16S rRNA序列进行鉴定。收集L. animalis LGM培养液上清,与细菌脂多糖(LPS,2μg/mL)同时孵育Caco-2细胞24 h,体外研究L. animalis LGM对Caco-2细胞内Th细胞分化转录因子(T-bet,GATA3,ROR-γt和Foxp3) mRNA表达的影响;配制L. animalis LGM细菌悬液,研究L. animalis LGM灌胃对DSS诱导结肠炎小鼠症状及结肠Th细胞分化转录因子和细胞因子(IFN-γ,IL-4,IL-17和IL-10) mRNA表达的影响,表达结果采用荧光定量PCR法检测。【结果】与对照组相比,L.animalisLGM培养液上清显著上调Caco-2细胞内ROR-γt与Foxp3 mRNA表达(P0.05),显著下调GATA3、IL-4、IL-17和TGF-βmRNA表达(P0.05)。L. animalis LGM灌胃显著上调小鼠结肠内ROR-γt和Foxp3的表达(P0.05),显著降低了促炎因子IL-4和IL-17的表达(P0.05),阻止了小鼠结肠长度缩短(P0.05)。【结论】猪肠道分离L. animalis LGM表现出对Th细胞分化转录因子的选择性调节,显著上调Caco-2细胞及结肠炎小鼠ROR-γt与Foxp3mRNA表达。降低DSS诱导结肠炎小鼠炎症水平,对DSS诱导结肠炎起保护作用,有助于维护肠道环境稳态。     

一株食源性蜡样芽孢杆菌的分离及其对小鼠肠黏膜免疫相关因子和肠道菌群的影响

蜡样芽孢杆菌(Bacillus cereus)是一种常见的食源性致病菌,误食被其污染的食品会引起呕吐或者腹泻,严重者还会死亡。本研究利用划线培养法从腐败米饭中分离培养出一株蜡样芽孢杆菌。通过药敏试验分析及毒力相关基因PCR扩增对其致病性及耐药性进行分析,并将细菌培养物经过腹腔注射进行小鼠感染试验,检测其对小鼠肠黏膜免疫相关因子和肠道菌群的影响,从而分析该病原菌的致病途径及防治手段。结果显示,本研究所分离的食源性蜡样芽孢杆菌,对诺氟沙星、呋喃妥因、四环素、米诺环素、环丙沙星、大观霉素、克林霉素、红霉素、克拉霉素、氯霉素、左氟沙星和万古霉素敏感,对复方新诺明、苯唑西林和青霉素G具有耐受性。该菌株携带hblA、hblC、hblD、nheA、nheB、nheC和entFM 7种毒力相关基因,可产生致腹泻型毒素。小鼠感染后,出现腹泻,肠黏膜免疫球蛋白和炎性因子表达水平显著上调;肠道菌群检测结果表明,经该蜡样芽孢杆菌感染后,小鼠肠道菌群组成发生变化,标志着机体健康的拟杆菌门(Bacteroidetes)中的uncultured_bacterium_f_Muribaculaceae丰度显著下降,而标志着菌群失调的变形菌门(Proteobacteria)条件致病菌uncultured_bacterium_f_Enterobacteriaceae丰度显著升高,且与IgM和IgG呈显著正相关(P<0.05)。这些研究结果表明,携带致腹泻型毒力相关基因的致病性蜡样芽孢杆菌感染机体后,可以通过影响肠道菌群激活免疫系统。     

双歧杆菌对SD大鼠空肠黏膜分泌型免疫球蛋白A的影响

目的研究双歧杆菌的免疫佐剂效应。方法将SD大鼠随机分成3组,即卵清白蛋白（OVA）组,双歧杆菌＋OVA组和PBS空白对照组。各组给予免疫后,收集长约3cm的空肠,通过免疫组织化学技术和Qwin图像处理系统,对空肠分泌型免疫球蛋白A（sIgA）的定位进行系统分析,研究同时注射OVA和双歧杆菌后对SD大鼠在不同时期空肠中sIgA阳性细胞分泌的影响。结果在免疫后的SD大鼠的空肠内均产生了特异性的分泌型抗体sIgA,其中双歧杆菌＋OVA组显著高于其他2组（P〈0．05）。结论双歧杆菌经胃肠道黏膜免疫可诱导有效的免疫佐剂效应。     

免疫共沉淀联合质谱对肝细胞核因子3β蛋白复合体的分离鉴定

目的:利用免疫共沉淀联合质谱分析方法,在人肝癌细胞系细胞株HepG2内筛选与肝细胞核因子(HNF)3β相互作用的蛋白质。方法:通过免疫共沉淀,用特异抗体分离出HNF3β蛋白复合体,质谱鉴定其成分,NCBI数据库检索各个蛋白质的功能,分析筛选与HNF3β相互作用的蛋白质。结果:利用免疫共沉淀联合质谱分析筛选到32个与HNF3β相互作用的候选蛋白质,数据库检索发现LMNA与HNF3β具有相同功能注释,参与转录调控。结论:在细胞内HNF3β与多种蛋白质有相互作用,协同参与转录调控,推测HNF3β/LMNA之间相互作用的变化与葡萄糖及脂肪代谢及相关疾病的发生有关。     

表达分泌型脑源性神经营养因子的腺相关病毒载体的构建及其表达的检测

采用PCR和体外连接的方法构建了带有信号肽的脑源性神经营养因子(BDNF)的融合基因, 将此基因插入到腺相关病毒载体穿梭质粒pSNAV, 然后用重组质粒pSNAV-Ig-BDNF转染包装细胞, 筛选出永久细胞株后, 用携带腺相关病毒rep及cap基因的单纯疱疹病毒超感染包装细胞, 成功包装出含有目的基因的一型血清型腺相关病毒。经PCR鉴定该病毒含有目的基因片段, 被该病毒感染的细胞裂解液经Western blotting 证实有BDNF表达, 其培养液经ELISA检测证实含有高水平的BDNF蛋白。该病毒与小量的非复制型腺病毒Ad-null联合应用时, 其BDNF蛋白的表达水平显著提高。可弥补腺相关病毒起效慢、对细胞转导效率低的弱点, 为今后应用AAV作基因治疗提供了实验证据。     

表达分泌型脑源性神经营养因子的腺相关病毒载体的构建及其表达的检测

采用PCR和体外连接的方法构建了带有信号肽的脑源性神经营养因子(BDNF)的融合基因, 将此基因插入到腺相关病毒载体穿梭质粒pSNAV, 然后用重组质粒pSNAV-Ig-BDNF转染包装细胞, 筛选出永久细胞株后, 用携带腺相关病毒rep及cap基因的单纯疱疹病毒超感染包装细胞, 成功包装出含有目的基因的一型血清型腺相关病毒。经PCR鉴定该病毒含有目的基因片段, 被该病毒感染的细胞裂解液经Western blotting 证实有BDNF表达, 其培养液经ELISA检测证实含有高水平的BDNF蛋白。该病毒与小量的非复制型腺病毒Ad-null联合应用时, 其BDNF蛋白的表达水平显著提高。可弥补腺相关病毒起效慢、对细胞转导效率低的弱点, 为今后应用AAV作基因治疗提供了实验证据。     

益生菌对肠易激综合征患者肠道微环境及免疫功能的影响

目的:探讨益生菌对肠易激综合征患者肠道微环境及免疫功能的影响,为临床相关研究提供参考。方法:选择2013年6月-2015年6月在我院接受治疗的肠易激综合征患者79例作为研究对象,根据治疗方案不同分为研究组(47例)和对照组(42例)。研究组患者采用双歧三联活菌联合美沙拉嗪治疗,对照组患者给予美沙拉嗪治疗。观察并比较两组患者治疗前后肠道内肠杆菌、双歧杆菌、乳酸杆菌、粪肠球菌及类杆菌的变化情况,以及血清CD4~+,CD8~+及CD4~+/CD8~+水平,并评价两组的临床疗效。结果:治疗前,两组患者肠道菌群数量比较,差异无统计学意义(P0.05);治疗后,两组患者肠道内大肠杆菌及粪肠球菌数量均减少,且研究组少于对照组,差异具有统计学意义(P0.05);治疗后,两组患者肠道内乳酸杆菌、双歧杆菌和类杆菌数量均增加,且研究组高于对照组,差异具有统计学意义(P0.05)。治疗前,两组患者免疫功能指标比较,差异无统计学意义(P0.05);治疗后,两组患者CD4~+T细胞以及CD4~+/CD8~+均显著高于治疗前,而CD8~+T细胞显著降低,差异具有统计学意义(P0.05);治疗后,研究组患者CD4~+T细胞及CD4~+/CD8~+均高于对照组,而CD8+T细胞低于对照组,差异具有统计学意义(P0.05)。研究组患者的临床疗效(91.67%)高于对照组(76.67%),但两组比较,差异无统计学意义(P0.05)。结论:益生菌制剂治疗不仅能够调节肠易激综合征患者肠道内的菌群平衡,而且能够改善患者的免疫功能,值得临床推广应用。     

球囊菌侵染对意大利蜜蜂幼虫肠道免疫与解毒相关基因表达及蛋白质、脂质和糖类含量的影响

[目的]球囊菌Ascosphaera apis是一种真菌性病原体,可以侵染意大利蜜蜂Apis mellifera ligustica幼虫,导致蜂群患白垩病.本研究通过球囊菌侵染意大利蜜蜂幼虫,探究对意大利蜜蜂幼虫肠道免疫解毒相关基因表达及蛋白质、脂质和糖类含量的影响,分析球囊菌侵染胁迫下意大利蜜蜂免疫应答反应与营养物质...     

Fermentation of coconut water by probiotic strains Lactobacillus acidophilus L10 and Lactobacillus casei L26

Coconut water is becoming an increasingly popular beverage and sports drink in tropical countries due to its high mineral content. Probiotic fermentation of coconut water would provide consumers with a novel probiotic beverage which can provide both hydration and probiotic benefits. The aim of this study was to assess the growth, survival and fermentation performance of two probiotic bacteria in coconut water. Lactobacillus acidophilus L10 and L. casei L26 grew well in coconut water and showed similar growth patterns. The viable cell count of the two probiotic cultures reached approximately 10⁸ CFU/ml after 2 days fermentation at 37 °C and maintained approximately10⁷–10⁸ CFU/ml after 26 days at 4 °C. Changes in total soluble solids (^oBrix), pH, sugars, organic acids and minerals were similar between the two probiotic cultures, except for fructose, glucose, copper, phosphorus and lactic, acetic and malic acids. There were significant variations between the two cultures in their ability to produce and consume these compounds. L. acidophilus produced higher amounts of 2-heptanone, 2-nonanone, benzaldehyde, 2-heptanol, 2-nonanol, δ-octalactone and δ-dodecalactone, whereas L. casei produced higher amounts of acetic acid, diacetyl, acetoin, δ-decalactone, 3-methyl-3-buten-1-ol, linalool, 1-octanol, p-tolualdehyde and ethyl 2-hydroxypropanoate. There was no substantial change in mineral content. These results suggest the feasibility of fermenting coconut water into a probiotic beverage, especially for sports nutrition, with the dual benefits of electrolytes and probiotics.     

Functional Analysis of Putative Adhesion Factors in Lactobacillus acidophilus NCFM

Lactobacilli are major inhabitants of the normal microflora of the gastrointestinal tract, and some select species have been used extensively as probiotic cultures. One potentially important property of these organisms is their ability to interact with epithelial cells in the intestinal tract, which may promote retention and host-bacterial communication. However, the mechanisms by which they attach to intestinal epithelial cells are unknown. The objective of this study was to investigate cell surface proteins in Lactobacillus acidophilus that may promote attachment to intestinal tissues. Using genome sequence data, predicted open reading frames were searched against known protein and protein motif databases to identify four proteins potentially involved in adhesion to epithelial cells. Homologous recombination was used to construct isogenic mutations in genes encoding a mucin-binding protein, a fibronectin-binding protein, a surface layer protein, and two streptococcal R28 homologs. The abilities of the mutants to adhere to intestinal epithelial cells were then evaluated in vitro. Each strain was screened on Caco-2 cells, which differentiate and express markers characteristic of normal small-intestine cells. A significant decrease in adhesion was observed in the fibronectin-binding protein mutant (76%) and the mucin-binding protein mutant (65%). A surface layer protein mutant also showed reduction in adhesion ability (84%), but the effect of this mutation is likely due to the loss of multiple surface proteins that may be embedded in the S-layer. This study demonstrated that multiple cell surface proteins in L. acidophilus NCFM can individually contribute to the organism's ability to attach to intestinal cells in vitro.     

嗜酸乳杆菌固态发酵的初步研究

研究了嗜酸乳杆菌在 4种不同固态基质 (豆渣、麦麸、芋艿渣、纤维素 )中的生长情况 ,结果显示 :1)豆渣作为基质时嗜酸乳杆菌生长最佳 ,最高活菌数可达 1.6 45× 10 9cfu/g ,麦麸次之 ,达 3.2× 10 8cfu/g ,纤维素最差 ,仅为 2 .1× 10 7cfu/g ,并且豆渣固态培养基优于对照的 3种液体培养基 ;2 )豆渣的含水量以 75 %左右为宜 ;3)通过加入可中和所产酸量的CaCO3 ,并用缓冲液代替水配制培养基 ,结果发现用pH 6 .0的缓冲液配制的培养基 ,培养后活菌量可达 2 .12 4× 10 9cfu/g ,,比对照用自来水提高了 4.2倍 .     

Effects of Probiotic Lactobacillus acidophilus Strain INMIA 9602 Er 317/402 and Putative Probiotic Lactobacilli on DNA Damages in the Small Intestine of Wistar Rats In Vivo

Probiotics and Antimicrobial Proteins - Double-strand breaks in the DNA of the small intestine in male Wistar rats were studied using a neutral comet assay after 7&nbsp;days of feeding with a...     

Viable Intestinal Passage of a Canine Jejunal Commensal Strain Lactobacillus acidophilus LAB20 in Dogs

The strain Lactobacillus acidophilus LAB20 with immunomodulatory properties was previously found dominant in the jejunal chyme of four dogs, and the novel surface layer protein of LAB20 suggested its competitive colonization in canine gut. To evaluate the persistence and survival of LAB20 in healthy dogs, LAB20 was fed to five healthy pet dogs for 3 days, at a dosage of 10⁸ CFU daily as fermented milk supplement. The fecal samples, from 1 day prior to feeding, three continuous feeding days, and on day 5, 7, 14, and 21, were collected for strain-specific detection of LAB20 using real-time PCR. We found that LAB20 count was significantly increased in dog fecal samples at the second feeding day, but rapidly decreased after feeding ceased. The fecal samples from prior to feeding, during feeding, and post-cessation days were plated onto mLBS7 agar, from where LAB20 was recovered and distinguishable from other fecal lactobacilli based on its colony morphotype. Using strain-specific PCR detection, the colonies were further verified as LAB20 indicating that LAB20 can survive through the passage of the canine intestine. This study suggested that canine-derived strain LAB20 maintained at high numbers during feeding, viably transited through the dog gut, and could be identified based on its colony morphotype.