基于16S rRNA高通量测序技术分析小鼠实验性结肠炎肠道菌群结构特征

目的:明确溃疡性结肠炎(ulcerative colitis,UC)与肠道菌群间的相互关系,为寻找简单、安全的UC治疗方法提供实验基础。方法:通过3%硫酸葡聚糖钠(dextran sulfate sodium,DSS)诱导建立小鼠溃疡性结肠炎模型,观察检测体重变化、肠道病理改变,采用16S rRNA高通量测序技术,检测小鼠粪便中的肠道菌群,并分析比较结肠炎小鼠与正常小鼠间肠道菌群的多样性及丰度。同时,利用已有数据库,预估肠道菌群内的功能基因构成,分析2组间的菌群基因功能差异。结果:溃疡性结肠炎小鼠体重明显减轻,且肠道上皮完整性被破坏,同时肠道菌群多样性明显降低,溃疡性肠炎组小鼠肠道内双歧杆菌属降至0.2%明显低于正常组小鼠(P0.05),而乳酸杆菌属平均丰度也显著降低至2.9%(P0.05),提示可能是通过影响机体代谢从而造成溃疡性结肠炎的发生、发展。结论:溃疡性结肠炎小鼠的肠道菌群多样性降低,菌群分布均发生显著改变。     

速激肽受体2对小鼠溃疡性结肠炎的影响

目的: 利用速激肽受体2（tachykinin receptor 2,Tacr2）基因敲除小鼠及诱导溃疡性结肠炎（ulcerative colitis,UC）小鼠模型,探讨Tacr2在小鼠UC发生、发展中的作用。方法: 小鼠按照基因型分成野生型组和基因敲除（纯合子）组,再按照给药与否进一步分成野生型造模组、纯合子造模组、野生型空白组、纯合子空白组。给造模组小鼠口服右旋葡聚糖硫酸钠（dextran sodium sulfate, DSS）构建UC小鼠模型,观察其UC活动指数及结肠组织中病理学改变、炎症因子及上游转录因子核因子NF-κB（nuclear factor-kappa B,NF-κB）的变化。结果: Tacr2基因敲除的纯合子小鼠经DSS诱导的UC症状比野生型造模组小鼠明显加重。进一步分析基础状态下结肠的免疫活性,发现在基础状态下,纯合子小鼠与野生型小鼠相比,结肠免疫活性明显降低,表现为结肠黏膜中 IL-1β、TNF-α、IL-6和NF-κB水平降低。结论: Tacr2对UC的发生、发展有抑制作用。     

联用乳酸杆菌和丁酸梭菌对小鼠急性溃疡性结肠炎的影响

目的观察联用乳酸杆菌和丁酸梭菌对小鼠急性溃疡性结肠炎的疗效并探讨其治疗机制。方法建立DSS诱导的小鼠急性溃疡性结肠炎（UC）模型,观察给予乳酸杆菌和丁酸梭菌治疗后,小鼠结肠黏膜的病理改变和TNF—α和组织因子（TF）表达的变化。结果乳酸杆菌和丁酸梭菌可明显减轻小鼠结肠黏膜的损伤;可明显抑制TNF—α和TF的表达,尤以两菌合用组抑制作用最强。结论乳酸杆菌和丁酸梭菌对DSS诱导的UC有治疗作用,对TNF-α和TF表达的协同抑制作用可能足其发挥协调治疗作用的分子机制。     

联用乳酸杆菌和丁酸梭菌治疗小鼠急性溃疡性结肠炎的实验研究

摘要 目的：观察联用乳酸杆菌和丁酸梭菌对小鼠急性溃疡性结肠炎的疗效并探讨其治疗机制。方法：建立DSS诱导的小鼠急性UC模型,观察给予乳酸杆菌和丁酸梭菌治疗后,小鼠结肠粘膜的病理改变和TNF-α和P-SEL表达的变化。结果：乳酸杆菌和丁酸梭菌可明显减轻小鼠结肠粘膜的损伤;可明显抑制TNF-α和P-SEL的表达,尤以两菌合用组抑制作用最强。结论：乳酸杆菌和丁酸梭菌对DSS诱导的UC有治疗作用,对TNF-α和P-SEL表达的协同抑制作用可能是其发挥协调治疗作用的分子机制。     

NEC患儿肠道菌群的优势菌属及益生菌制剂疗效分析

目的探讨新生儿坏死性小肠结肠炎(NEC)患儿肠道菌群的优势菌属及益生菌制剂的疗效。方法选取2018年2月至2020年2月于该院治疗的30例NEC患儿作为观察组,同时选取于该院体检的健康儿童30例作为对照组。比较观察组和对照组儿童肠道菌群的丰富度并分析两组儿童肠道菌群的优势菌属。采用随机数字表法,将观察组患儿分为A组及B组;A组患儿采用酪酸梭菌二联活菌散剂联合奥曲肽治疗,B组患儿仅采用奥曲肽治疗;两组患儿均治疗1周,比较两组患儿的治疗效果以及肠道菌群的差异。结果观察组肠道菌群丰富度显著低于对照组(P0.05),两组儿童的肠道细菌优势菌属条带相似度接近100%。观察组患儿双歧杆菌及大肠埃希菌呈下降趋势。经1周治疗,A组治疗总有效率显著高于B组(P0.05);治疗后,A、B两组患儿的粪便乳酸杆菌、双歧杆菌、肠球菌、真杆菌菌落数均有所升高(P0.05),且A组患儿乳酸杆菌、双歧杆菌、肠球菌、真杆菌菌落数明显高于B组。结论 NEC患儿肠道菌群中的双歧杆菌以及大肠埃希菌呈下降趋势,可通过采用益生菌制剂进行辅助治疗。     

溃疡性结肠炎患者及其配偶的肠道菌群构成与多样性分析

目的比较同一生活环境下溃疡性结肠炎(UC)患者与其配偶(UF),及健康人群的肠道菌群差异,探讨生活环境对UC患者肠道菌群的影响。方法招募UC患者8例、同一生活坏境下患者配偶8例及普通健康人群8例,分为UC组、UC配偶组(UF组)、健康人群组(ZC组)。分别收集三组人群的粪便样本,通过16SrRNA基因测序技术进行菌群分析,比较三组样本间肠道菌群的差异性。结果肠道菌群多样性分析表明三组间菌群丰富度及群落多样性差异无统计学意义(P>0.05)。肠道菌群构成分析表明在属分类水平上,ZC组及UF组中考拉杆菌属的相对丰度比UC组增高(P<0.05);ZC组及UC组中锥形杆菌属的相对丰度比UF组降低(P<0.05);UC组及UF组中奥尔森菌属、霍尔德曼菌属、克雷伯菌属的相对丰度比ZC组增高(P<0.05)。结论考拉杆菌属、奥尔森菌属及克雷伯菌属可能是预测UC肠道菌群失调及疾病预后的生物学标志物。生活环境会影响UC患者肠道菌群变化,但并不是UC发病的决定性因素。     

二甲双胍与美沙拉嗪在小鼠结肠炎中的疗效比较

目的 探讨二甲双胍与美沙拉嗪在治疗小鼠结肠炎中的疗效差异性。方法 选取6 ~ 8周龄的C57BL/6雄性小鼠32只,随机分为Control组、硫酸葡聚糖（DSS）组、DSS+美沙拉嗪组、DSS+二甲双胍组,每组8只。Control组小鼠自由饮用高压灭菌双蒸水,并每日予0.2 ml 磷酸盐缓冲液（PBS）灌胃。采用3%DSS诱导C57BL/6雄性小鼠结肠炎模型。DSS组小鼠自由饮用3%DSS溶液,每日予0.2 ml PBS灌胃,DSS+美沙拉嗪组、DSS+二甲双胍组自由饮用3%DSS溶液,予100 mg/（kg·d） 的美沙拉嗪或100 mg/（kg·d） 二甲双胍灌胃。评估各组小鼠体质量变化、结肠长度、疾病活动指数、组织病理学改变及肠道炎症因子表达情况。结果 与DSS组相比,DSS+二甲双胍与DSS+美沙拉嗪组均可降低小鼠结肠炎所致的体质量丢失、疾病活动度评分、组织病理学评分及肠道炎症因子表达水平（P均< 0.05）,但DSS+二甲双胍组与DSS+美沙拉嗪组上述指标比较差异均无统计学意义（P均> 0.05）。结论 二甲双胍可以减轻DSS诱导的小鼠结肠炎,其疗效不亚于美沙拉嗪。     

严重烧伤伴腹泻患者口服双歧杆菌制剂的疗效观察及护理

目的：观察双歧杆菌制剂治疗严重烧伤病人腹泻的作用。方法：选取烧伤后腹泻病人26例，从腹泻当天开始喂服双歧杆菌活菌制剂，于喂服前及喂服后第3天及第6天采集粪便做粪便菌群分析。另选10名健康成年人粪便标本为对照组。结果：严重烧伤后粪便菌群总菌量较正常显下降，肠杆菌轻度下降，双歧杆菌下降近1000倍，酵母样真菌显升高，说明肠道菌群发生紊乱。用双歧杆菌制剂治疗6d后腹泻均停止，肠道菌群趋向正常，与对照组和治疗前比较均存在显性差异（P＜0．05或P＜0．01）。结论：口服双歧杆菌制剂能调整肠道菌群紊乱，起到治疗腹泻的作用。注意饭后给药，低温（4℃）存放药物。     

青春双歧杆菌和嗜酸乳杆菌对小鼠急性实验性结肠炎的治疗作用

目的通过建立葡聚糖硫酸钠（DSS）诱导的急性期溃疡性结肠炎（UC）小鼠模型,探讨青春双歧杆菌和嗜酸乳杆菌诱导UC缓解的作用。方法 6～8周龄雌性BABL/c小鼠50只,体质量（20.0±2.0）g,随机分为5组：正常对照组（N-7组）、嗜酸乳杆菌（L-7组）、青春双歧杆菌组（B-7组）、柳氮磺胺吡啶（SASP）组（S-7组）和生理盐水对照组（NS-7组）,每组10只。除正常对照组外,采用自由饮用2.5%DSS（分子量3.6万～5万）7天建立小鼠UC模型,且从实验第1天开始,给予青春双歧杆菌、嗜酸乳杆菌（2×10^8 CFU/10 g）和SASP（15 mg/10 g）灌肠治疗7天,观察小鼠一般情况、粪便隐血,计算疾病活动指数（DAI）;实验结束时处死动物,测量全段结肠长度,行组织病理学检查并进行组织学评分。结果青春双歧杆菌、嗜酸乳酸杆均能诱导实验性结肠炎的缓解,其DAI积分、病理组织学评分均低于对照组（P〈0.05）;嗜酸乳酸杆的保护作用强于青春双歧杆菌和SASP。结论青春双歧杆菌和嗜酸乳酸杆菌能有效地减轻急性期UC炎症,且嗜酸乳酸杆菌更有利于诱导急性期UC的缓解。     

双歧杆菌对高脂饮食诱导的C57BL/6小鼠非酒精性脂肪肝的影响

目的： 研究双歧杆菌是否通过改变肠道菌群,减少肠道局部炎症,调节系统性炎症,从而减轻高脂饮食诱导的C57BL/6小鼠的非酒精性脂肪肝的发生发展。方法：采用雄性 C57BL/6 小鼠作为动物模型,分别给予对照饲料及高脂饲料喂养,同时予双歧杆菌干预。通过ELISA检测小鼠血清脂代谢及肝功能、TNF-α及LBP水平。通过油红O染色剂HE染色评估小鼠肝脏脂肪变性情况。通过real time PCR检测小鼠回肠、结肠、肝脏组织中炎症因子TNF-a、IL-6的表达。通过Illuminate Miseq平台微生物多样性测序检测小鼠粪便中肠道菌群的结构变化。应用SPSS19.0软件进行统计分析,多组间样本均数比较采用ANOVA进行检验,组间两两比较,p<0.05为差异具有统计学意义。结果：双歧杆菌干预组C57BL/6小鼠肝脏脂肪变程度明显低于高脂饲料喂养组。与高脂肪饲料喂养组相比,双歧杆菌干预组炎症因子TNF-a、IL-6表达明显下降,肠道炎症减轻。与对照组小鼠相比,高脂饲料诱导的NAFLD小鼠肠道菌群物种丰度和多样性减少,存在肠道菌群改变,PcoA分析(主成分分析)及NMDS(非度量尺度分析)结果显示疾病模型组与阴性对照组肠道菌群构成分隔较远,构成上存在显著差异。菌属差异分析表现为高脂组拟杆菌门增多,厚壁菌门减少,拟杆菌属增多。     

前列腺素E1治疗溃疡性结肠炎疗效观察

目的观察前列腺素E1对溃疡性结肠炎患者的治疗效果。方法反复发作的活动期轻、中度溃疡性结肠炎患者35例，随机分为2组：治疗组口服柳氮磺吡啶1g，3次／d；并应用前列腺素E1 100μg／d静脉滴注，疗程2周。对照组仅服柳氮磺吡啶1g，3次／d。观察患者疾病活动指数（DAI）及血沉（ESR）、C-反应蛋白（CLIP）、血小板计数（BPC）等的变化。结果治疗组72．2％（13／18）临床病情缓解，对照组29．4％（5／17）缓解（P〈0．05）。治疗组DAI、ESR、CPR、BPC等的改善程度与对照组比较差异均有显著性（均为P〈0．05）。两组均未发现明显不良反应。结论溃疡性结肠炎患者应用前列腺素E1治疗可取得满意的效果。     

Probiotics in inflammatory bowel disease: controlled trials and perspectives

Probiotics may modulate intestinal flora and immunity and are therefore studied in an attempt to modulate experimental colitis or human inflammatory bowel disease. We analysed randomised controlled trials performed using probiotics in humans with Crohn's disease, ulcerative colitis and pouchitis. Perspectives include the use of genetically modified micro-organisms to deliver anti-inflammatory agents to the gastrointestinal tract.     

Effect of ceftazidime and gentamicin on the oropharyngeal and faecal flora of patients with haematological malignancies

Thirty-four patients with haematological malignancies were studied to investigate the effect of empirical broad-spectrum antibiotic therapy (ceftazidime and gentamicin) on the gastro-intestinal flora. Twenty-five patients with acute myeloid leukaemia or post-autologous bone-marrow transplantation were given framycetin, nystatin and colistin (Fracon), and two patients with non-Hodgkin's Lymphoma were on co-trimoxazole, as long-term gut prophylaxis. Semi-quantitative microbiology was carried out on oropharyngeal swabs and quantitative microbiology on faecal specimens. The oropharyngeal flora consisted mainly of streptococci, coagulase-negative staphylococci and coryneforms, and was little affected by ceftazidime/gentamicin. A strain of Enterobacter cloacae resistant to ceftazidime and gentamicin colonized one patient, who later developed septicaemia. The faecal flora of patients on Fracon was dominated by enterococci; the few enterobacteria present were eliminated by ceftazidime/gentamicin. The anaerobic flora was absent in 15% of patients; in the remainder, it consisted mainly of Bacteroides spp., and was little affected by ceftazidime/gentamicin. The faecal flora of patients not on Fracon always contained anaerobes, and some strains of enterobacteria persisted throughout antibiotic treatment. None of the patients was colonized by Clostridium difficile or Pseudomonas aeruginosa. Broad-spectrum therapy with ceftazidime and gentamicin appeared to have little effect on the gastro-intestinal flora, except to encourage the overgrowth of enterococci and reduce the numbers of enterobacteria.     

复方灌肠液治疗溃疡性结肠炎的临床观察

目的观察西医内科基础治疗辅以复方灌肠液保留灌肠治疗溃疡性结肠炎的临床疗效。方法将80例患者随机分为治疗组及对照组各40例,对照组口服柳氮磺胺毗啶（SASP）1．0g,4次／d;治疗组在此基础上加用复方灌肠液保留灌肠,14d为1疗程,连用3个疗程。比较两组患者治疗前后的临床症状、疾病活动指数（DAI）、肠镜表现、复发情况及不良反应。结果治疗组患者的临床症状、DAI、肠镜表现明显改善,复发率低,对照组改善不明显,差异有统计学意义（P〈0．05）,两组不良反应发生率差异无统计学意义（P〉0．05）。结论SASP口服联合复方灌肠液保留灌肠治疗溃疡性结肠炎,疗效显著,简便实用,值得临床推广。     

Effect of trimethoprim on the occurrence of drug-resistant coliform bacteria in the faecal flora.

The occurrence of drug-resistant coliform bacteria was studied in the faecal flora of 30 persons receiving for 3 weeks either trimethoprim alone, a combination of sulphamethoxazole and trimethoprim, or a combination of sulphamethoxydiazine and sulphamethoxazole. Bacterial sensitivity was tested against trimethoprim, sulphamethoxazole-trimethoprim, sulphamethoxazole, and sulphaisodimidine. After treatment with trimethoprim alone, no increase in the occurrence of strains resistant to either trimethoprim or sulphonamides was observed. After treatment with sulphamethoxazole-trimethoprim, the faecal flora contained an increased percentage of sulphonamide-resistant coliforms but significantly less than found after treatment with sulphamethoxydiazine-sulphamethoxazole. In the persons receiving the sulphonamides only, a rapid increase in sulphonamide-resistant coliforms was observed. During the whole study, only one trimethoprim-resistant coliform strain was detected.     

Infliximab for the treatment of pouchitis

Pouchitis is not a rare complication that develops after an ileal-pouch anastomosis, performed after colectomy in patients refractory to treatment or with complicated ulcerative colitis. This condition may become chronic and unresponsive to medical therapies, including corticosteroids, antibiotics and probiotics. The advent of biological therapies (tumor necrosis factor-α inhibitors) has changed the course of these complications. In particular, in these cases, infliximab (IFX) may represent a safe and effective therapy in order to avoid the subsequent operation for a permanent ileostomy. This article reviews the therapeutic effects of one of the most widely used anti-tumor necrosis factor-α molecules, IFX, for the treatment of complicated pouchitis (refractory to conventional treatment and/or fistulizing).     

久泻灵冲剂对溃疡性结肠炎大鼠GSH-Px、SOD及MDA影响的实验研究

目的观察久泻灵冲剂对溃疡性结肠炎大鼠模型的影响,为临床用药的有效性提供实验依据。方法采用异种异体结肠黏膜组织致敏法和乙酸局部灌肠相结合的方法制造动物模型。将Wistar大鼠随机分为空白对照组、模型对照组、阳性对照组及久泻灵冲剂大、中、小剂量组。连续给药14 d,末次给药后24 h处死大鼠,制备结肠组织匀浆,生化法检测结肠组织中GSH-Px、SOD、MDA的含量,结肠组织HE染色后进行病理学组织评分。结果久泻灵可明显降低结肠组织中MDA的活性水平,升高GSH-Px、SOD的活性,降低病理组织学评分。结论久泻灵冲剂对溃疡性结肠炎模型大鼠有显著的干预作用,其机制可能与抗氧自由基损伤有关。     

Probiotics modulate inflammatory cytokine secretion from inflamed mucosa in active ulcerative colitis

Enteric microflora of ulcerative colitis patients becomes aberrant. The abnormal interaction between microflora and intestinal mucosal immune system leads the mucosal inflammation. Probiotic administration may recover the commensal microflora and normalise the host-microbial interaction. In this experiment, we cocultured colonic biopsies from active ulcerative colitis patients with bifidobacterium to investigate the modulation effect of probiotics on inflamed colonic tissues and its possible mechanism. Colonic biopsies from active ulcerative colitis were cocultured for 24 h with Bifidobacterium longum. Tumour necrosis factor (TNF)-alpha and interleukin (IL)-8 in supernatants were measured using enzyme-linked immunosorbent assays, the biopsies were fixed using paraffin and the expression of NF-kappaB P65 of tissues was studied using immunohistochemical staining. The concentrations of TNF-alpha and IL-8 in supernatants of tissues cocultured with probiotics were lower than those cultured alone. The number of lamina propria mononuclear cells (LPMC) with nuclear factor-kappa B (NF-kappaB) P65 positive in cocultured tissues was also decreased. When cocultured with inflamed tissues of active ulcerative colitis, probiotics could inhibit NF-kappaB activation in LPMC and down-regulate inflammatory cytokine secretion from inflamed tissues of active ulcerative colitis.