利用生物发光活体成像技术观察肠黏膜损伤状态下细菌易位的初步研究

[目的]临床上各种疾病会导致肠黏膜屏障损伤并伴随大量的细菌易位发生。本实验拟使用发光细菌的小鼠活体成像技术,观察肠黏膜损伤状态下易位细菌的动态移位过程。[方法]构建重组荧光脂酶基因(Lux)的质粒pXen-1、pXen-18,转化大肠杆菌DH5a,筛选阳性细菌克隆,扩增培养后,定量(107)注射到肠黏膜屏障损伤模型小鼠肠腔不同部位内,利用生物发光活体成像技术进行细菌易位的病理生理观察研究。实验分为四组:小肠注射菌实验组和对照组、盲肠注射菌实验组和对照组。其中实验组为肠道缺血再灌注损伤,对照组无任何干预措施。[结果]发光细菌小鼠活体成像观察到,存在肠黏膜损伤的小鼠肠腔内的细菌数量及分布发生明显的改变,提示细菌易位在肠黏膜损伤时起着重要的病理生理作用。注入小肠内的发光细菌数量在试验过程中明显下降,提示小肠肠腔环境可以杀灭一定的细菌。[结论]生物发光活体成像技术,可以直观、可靠、敏感地观察肠道内细菌的存活状态,并可观察肠黏膜损伤状态下细菌易位的动态过程。该技术为肠黏膜屏障损伤细菌易位机制的研究提供了一个有力的工具。     

人类免疫缺陷病毒感染相关肠屏障损伤及其靶向治疗的研究进展

高效抗逆转录病毒治疗(HAART)能有效地抑制人类免疫缺陷病毒(HIV)复制,大大降低获得性免疫缺陷综合征(艾滋病,AIDS)的患病率和病死率,却无法彻底清除病毒,进而发展为慢性病毒感染性疾病。慢性HIV感染破坏宿主免疫系统导致肠屏障破坏、肠黏膜功能紊乱及菌群易位加快疾病进程。而重建肠道微生态平衡和改善肠黏膜功能对重建宿主免疫系统至关重要。以下综述近年来关于艾滋病相关肠屏障损伤及其靶向治疗的研究进展,为进一步研究和改进艾滋病治疗措施、降低HIV感染患病率和病死率提供参考。     

HIV感染期间持续性免疫激活与肠道微生物易位关系的研究

由人类获得性免疫缺陷病毒(HIV)感染引起的病理学改变是复杂、多方面的,其中感染过程中导致免疫激活和疾病进展的一个关键因素是微生物易位,在胃肠道表现较早,并出现相应的临床症状。人类和动物感染SIV后,导致细菌易位形成的主要因素包括胃肠黏膜上皮屏障破坏和持续性免疫激活。以下就HIV感染期间持续性免疫激活与肠道微生物易位的关系进行综述。     

多黏菌素E诱导肠道菌群失调对肠屏障功能和细菌易位的影响

目的:应用多黏菌素E给小鼠灌胃诱导建立肠道菌群失调动物模型,研究肠道菌群失调对肠黏膜屏障和细菌易位的影响。方法:将20只小鼠随机分为两组,每组10只。实验组采用多黏菌素E按0.2 g/kg加入0.2 ml等渗盐水灌胃,1次/d,连续7 d。对照组用等剂量等渗盐水灌胃,1次/d,连续7 d。实验结束次日观察两组小鼠回肠黏膜病理形态、肠道菌群、回肠黏膜组织紧密连接(TJ)蛋白表达和器官(肝、脾、肾、淋巴结)细菌易位率等。结果:实验组小鼠盲肠黏膜和盲肠内容物的肠杆菌数量显著减少。常规病理检查发现,实验组小鼠回肠黏膜充血明显,绒毛稀疏,尖端有少量上皮坏死脱落,与对照组比损伤明显。透射电镜显示,实验组小鼠回肠上皮TJ的电子致密物质明显减少,TJ破坏。回肠黏膜组织中TJ蛋白Claudin-1、Occludin和ZO-1的表达显著下降,器官细菌易位率显著高于对照组。结论:多黏菌素E灌胃能够诱导肠道菌群失调,且导致肠屏障功能损伤和细菌易位。     

肠屏障损伤与肠源性感染

<正>肠源性感染(gut-derived infection)是严重创(烧)伤、大手术、失血性休克、重症急性胰腺炎等危重症病人死亡的主要原因之一~([1])。在上述病理条件下,肠道细菌可通过受损的肠黏膜侵入外周血和远隔器官,形成细菌易位,诱发全身性炎症反应、脓毒症和多器官功能障碍综合征(MODS),导致病人死亡~([2])。肠屏障(intestinal barrier)主要由肠上皮屏障、肠道微生物屏障和黏膜免疫屏障组成。以往细     

艾司洛尔对脓毒症大鼠肠黏膜屏障的影响

目的:探讨艾司洛尔对脓毒症大鼠肠黏膜屏障的保护作用. 方法:建立脓毒症大鼠模型,将大鼠随机分为对照组(脓毒症组)和艾司洛尔组(脓毒症模型+艾司洛尔组),每组8只.观察大鼠的基本生命体征、体外肠道的通透性和血清内毒素变化.同时,通过肝、脾和肠系膜淋巴结细菌培养观察细菌易位情况.最后,根据H-E染色结果评估大鼠肠道损伤. 结果:艾司洛尔组大鼠心率明显低于基础值(P＜0.05).与对照组比,艾司洛尔组大鼠细菌易位减少,体外肠道通透性降低.两组大鼠血清内毒素水平无显著性差异.艾司洛尔组大鼠肠道病理评分明显低于对照组. 结论:艾司洛尔能保护脓毒症大鼠肠黏膜屏障,减轻肠损伤.     

微生物酵素预防治疗肠道细菌易位的实验研究

目的探讨微生物酵素对他克莫司(FK506)作用下肠道细菌易位情况的影响。方法健康雄性Wistar大鼠84只,体重180～220g,随机分为对照组、免疫抑制组和预防治疗组;比较3组间肝和肠系膜淋巴结细菌培养阳性率、肠黏膜病理分析。结果免疫抑制组第5、7天脏器细菌培养阳性率分别为28.57%、42.86%,显著高于对照组的0、0(P<0.05);各时间点肠黏膜损伤评分均显著高于对照组(P<0.05);预防治疗组第7天脏器细菌培养阳性率为7.14%,显著低于免疫抑制组的0(P<0.05);第3、5、7天肠黏膜损伤评分均显著低于免疫抑制组(P<0.05)。结论 FK506可以引起肠黏膜上皮损伤,肠道机械屏障功能破坏,肠黏膜通透性增高,发生肠道细菌易位;应用微生物酵素在一定程度上能够减轻FK506引起的肠黏膜上皮损害,保护肠黏膜机械屏障功能,降低肠黏膜通透性,减少肠道细菌易位的发生。     

营养支持对肠道菌群与肠黏膜免疫的影响

肠内营养(EN)有助于保持肠黏膜的屏障功能,维护肠黏膜的结构完整性,促进肠黏膜固有免疫和获得性免疫的产生,维持肠道菌群的平衡和多样性。肠外营养(PN)能预防胃肠道功能障碍患者进一步营养不良的发生,挽救危重症患者的生命,然而PN会增加危重症患者感染并发症的发生。全肠外营养(TPN)时肠黏膜免疫受损,肠道菌群(GM)紊乱,滋养性喂养能显著改善肠黏膜的免疫功能,促进肠道内稳态的恢复。GM紊乱加重肠黏膜的炎症反应,或将进一步加重肠黏膜免疫的损害。因此,全面理解营养支持、GM与肠黏膜免疫之间的相互关系具有重要意义。     

条件必需氨基酸营养代谢与肠道健康

正常肠屏障功能的维持主要依赖于肠黏膜屏障.每天食人的氨基酸是肠黏膜的主要物质,也是合成肠道蛋白、一氧化氮和各种代谢产物的重要底物,具有维持肠道的完整性和生理功能.现就Gln、谷氨酸和精氨酸在肠道健康的营养和免疫的作用作一综述.     

肠道黏膜、肠道免疫和微生物群在肠源性白念珠菌感染中的作用

肠源性白念珠菌感染是指肠道内定植的白念珠菌在一定条件下发生易位,突破肠道,造成组织感染,甚至引发侵袭性白念珠菌感染。肠道黏膜作为念珠菌第一接触位点,是抵抗白念珠菌定植或侵入的第一道防线,常通过物理屏障和激活宿主免疫抑制感染。作为另一种防御机制,肠道内微生物群则通过调节pH, 分泌抗菌肽和竞争黏附点共同抵抗白念珠菌侵袭感染。本综述总结肠道黏膜、肠道免疫和微生物群这三个关键因素在肠源性白念珠菌感染中的作用, 为肠道定植引发侵袭性白念珠菌病的科学研究提供新思路。     

Impaired intestinal immunity and barrier function: a cause for enhanced bacterial translocation in alcohol intoxication and burn injury.

Alcohol intoxication is being recognized increasingly as the major factor in pathogenesis after burn injury. Findings from multiple studies support the suggestion that, in comparison with burn-injured patients who sustained injury in the absence of alcohol intoxication, burn-injured patients who sustained injury under the influence of alcohol exhibit higher rates of infection and are more likely to die. Thus, infection becomes the primary cause of death in burn-injured patients. Because the intestine is considered to be a major source of bacteria, studies in experimental animals have been designed to examine whether alcohol intoxication before burn injury enhances bacterial translocation from the intestine. Results of these studies have shown a several-fold increase in bacterial translocation from the intestine in the group of animals receiving combined insult of alcohol intoxication and burn injury compared with findings for the groups receiving either insult alone. Alcohol intoxication and burn injury independent of each other have also been shown to cause an increase in bacterial translocation. The gastrointestinal tract normally maintains a physical mucosal and immunologic barrier that provides an effective defense in keeping bacteria within the intestinal lumen. However, in injury conditions these defense mechanisms are impaired. Intestinal bacteria consequently gain access to extraintestinal sites. Intestine-derived bacteria are implicated in causing systemic infection and in subsequent multiple organ dysfunction in both immunocompromised patients and patients with injury, such as burn and trauma. In this article, we discuss three potential mechanisms that are likely to contribute to the increase in bacterial translocation in alcohol intoxication and burn injury: (1) increase in bacterial growth in the intestine, (2) physical disruption of mucosal barrier of the intestine, and (3) suppression of the immune defense in the intestine.     

乳杆菌对致病性大肠杆菌感染小鼠肠黏膜屏障功能的影响

目的:研究乳杆菌对致病性大肠杆菌感染小鼠肠黏膜屏障的保护作用.方法:将BALB/c小鼠随机分为对照组、致病性大肠杆菌感染组、乳杆菌JCM1081灌胃组、乳杆菌治疗组等,实验进行14 d.观察各组小鼠肠黏膜形态变化、肠道菌群变化、细菌移位以及肠黏膜钙黏蛋白表达的差异.结果:致病性大肠杆菌感染组小鼠与对照组相比,肠黏膜形态有明显改变,肠道内厌氧菌数量显著下降,肠杆菌和肠球菌数量明显增加,在肝、脾、肾以及肠系膜淋巴结中有细菌存在,但肠黏膜钙黏蛋白无明显变化.乳杆菌灌胃组小鼠各项指标无显著变化.乳杆菌治疗组小鼠各项指标变化程度较致病性大肠杆菌感染组显著减轻.结论:乳杆菌能黏附定植于肠黏膜并形成生物屏障,起着保护肠黏膜免受损伤的作用.     

益生菌在肠内营养中的应用及研究进展

防治肠源性感染的关键在于防止肠内细菌和(或)内毒素的移位,纠正肠内菌群平衡的紊乱和失调,改善肠黏膜屏障功能以及提高肠道局部的免疫功能。而益生菌是具有维持肠内菌群平衡、防止肠黏膜屏障的破坏、增强局部免疫功能、促进和改善消化道蠕动以及吸收等作用的非致病性微生物,可治疗和预防肠源性感染。以下综述益生菌在肠内营养的作用及研究进展。     

OK-432 reduces mortality and bacterial translocation in irradiated and granulocyte-colony stimulating factor (G-CSF)-treated mice

Bacterial translocation/Acute radiation syndrome/Endotoxin/G-CSF/OK-432 Acute radiation induces bacterial translocation from the gut, followed by systemic infection and sepsis. In order to reduce the mortality after acute whole body irradiation, it is essential to control bacterial translocation. In this study, we established a bacterial translocation assay as a sensitive method to detect minor mucosal injury by radiation. By utilizing this assay, we evaluated the adverse effects, if any, of hematopoietic reagents on the mucosal integrity in the respiratory and gastro-intestinal tracts. Bacterial translocation to the liver and spleen occurred after whole-body irradiation if the dose exceeded 6 Gy. The administration of G-CSF unexpectedly increased the bacterial translocation in 8 Gy-irradiated mice. The pharmaceutical preparation of low-virulent Streptococcus pyogenes, OK-432, significantly reduced the endotoxin levels in peripheral blood without any reduction of bacterial translocation. A combined treatment with G-CSF and OK-432 decreased bacterial translocation and prevented death. This result indicates that the early administration of G-CSF has an adverse effect on bacterial translocation, and that a combined treatment of G-CSF and OK-432 attenuates the adverse effect of G-CSF and improves the survival rate after acute irradiation.     

肠道粘膜屏障的损伤与保护

肠道依靠其机械、免疫屏障及正常的微生态环境等,能有效阻止肠道细菌移位和内毒素血症.但在创伤、应激等情况下,其结构和功能可受到严重损害,从而引起一系列病理生理的变化,充分认识肠粘膜屏障损伤的危害性,探讨其发生的机制及采取积极有效地保护措施,有助于防止疾病的发生和多器官功能障碍.     

The Multi-Component Causes of Late Neonatal Sepsis—Can We Regulate Them?

Elucidating the mechanisms of bacterial translocation is crucial for the prevention and treatment of neonatal sepsis. In the present study, we aimed to evaluate the potential of lactoferrin to inhibit the development of late-onset blood infection in neonates. Our investigation evaluates the role of key stress factors leading to the translocation of intestinal bacteria into the bloodstream and, consequently, the development of life-threatening sepsis. Three stress factors, namely weaning, intraperitoneal administration of Gram-positive cocci and oral intake of Gram-negative rods, were found to act synergistically. We developed a novel model of rat pups sepsis induced by bacterial translocation and observed the inhibition of this process by supplementation of various forms of lactoferrin: iron-depleted (apolactoferrin), iron-saturated (hololactoferrin) and manganese-saturated lactoferrin. Additionally, lactoferrin saturated with manganese significantly increases the Lactobacillus bacterial population, which contributes to the fortification of the intestinal barrier and inhibits the translocation phenomenon. The acquired knowledge can be used to limit the development of sepsis in newborns in hospital neonatal intensive care units.     

大黄和早期肠内营养对肠缺血-再灌注损伤大鼠肠屏障功能的影响

目的:观察大黄和早期肠内营养(EEN)对大鼠肠缺血-再灌注损伤(IRI)后肠黏膜屏障的影响.方法:将32只大鼠随机分为对照组、EN组、大黄组、大黄+EN组.在大鼠肠IRI模型上观察再灌注24h后肠黏膜形态学的变化、血清内毒素和细菌易位指标. 结果:肠IRI可导致肠黏膜发生严重损伤.肠IRI后24h,大黄组大鼠内毒素水平...     

Effects of probiotics on radiation-induced intestinal injury in rats

OBJECTIVE: Radiotherapy is an important aspect of multimodal cancer therapy, but radiation-induced acute intestinal injury is a common and serious problem. Disruption of morphologic mucosal integrity and normal bacterial microflora after abdominal radiation leads to malabsorption and bacterial translocation. METHODS: Lactobacillus bulgaricus strain isolated from yogurt was given as a probiotic to rats subjected to radiotherapy. On postradiation day 8 rats were killed. Mesenteric lymph nodes, liver, and spleen were excised for microbiologic examinations. Segments of jejunum, ileum, and colon were evaluated for the presence of inflammation, vascularity, and mucus cells. RESULTS: The results of this study suggest that probiotics may have a protective effect on intestinal mucosa. CONCLUSION: Probiotics added as substrates can be given by an oral or enteral route to patients who undergo radiotherapy to prevent radiation-induced enteritis and related malnutrition.     

Glutamine Supplementation Decreases Intestinal Permeability and Preserves Gut Mucosa Integrity in an Experimental Mouse Model

Background: Glutamine (GLN) is the preferred fuel for enterocytes, and GLN supplementation is critical during stressful conditions. The aim of this study was to evaluate the effect of GLN on intestinal barrier permeability and bacterial translocation in a murine experimental model. Methods: Swiss male mice (25–30 g) were randomized into 3 groups: (1) sham group; (2) intestinal obstruction (IO) group; (3) IO and GLN (500 mg/kg/d) group. Two different experiments were carried out to assess intestinal permeability and bacterial translocation. In the first experiment, the animals were divided into the 3 groups described above and received diethylenetriamine pentaacetate radiolabeled with technetium (^99mTc) on the eighth day. At different time points after intestinal obstruction, blood was collected to determine radioactivity. The animals were killed, and the small intestine was removed for histological analyses. In the bacterial translocation study, on the eighth day all groups received Escherichia coli labeled with ^99mTc. After 90 minutes, the animals underwent intestinal obstruction and were killed 18 hours later. Blood, mesenteric lymph nodes, liver, spleen, and lungs were removed to determine radioactivity. Statistical significance was considered when P ≤ .05. Results: The levels of intestinal permeability and bacterial translocation were higher in the IO group than in the sham and GLN groups (P < .05). GLN decreased intestinal permeability and bacterial translocation to physiologic levels in the treated animals and preserved intestinal barrier integrity. Conclusions: GLN had a positive impact on the intestinal barrier by reducing permeability and bacterial translocation to physiologic levels and preserving mucosal integrity.