Effect of pH on an in vitro model of gastric microbiota in enteral nutrition patients

Patients with dysphagia due to oropharyngeal disease or cerebrovascular accident require long-term nutritional support via enteral feeding, which often results in microbial overgrowth in the upper gastrointestinal (GI) tract. Gastric acid is the primary innate defense mechanism in the stomach and has been assumed to provide an effective barrier to microbial colonization at pH values of <4. To evaluate the efficacy of gastric acid as a barrier to overgrowth, the microbiota of gastric and duodenal aspirates was assessed by culturing methods. Additionally, a fermentor-based model incorporating enteral nutrition tubing of the gastric microbiota of enteral nutrition (EN) patients was constructed to assess the effect of pH on the microbiota. Results showed that gastric acidity had a relatively small effect on the numbers of microorganisms recovered from intestinal aspirates but did influence microbiota composition. Similarly, at pH 3 in the fermentor, a complex microbiota developed in the planktonic phase and in biofilms. The effect of pH on microbiota composition was similar in aspirates and in the fermentors. Candidas and lactobacilli were aciduric, while recoveries of Escherichia coli and Klebsiella pneumoniae decreased as pH was reduced, although both were still present in significant numbers at pH 3. Only Staphylococcus aureus and Bifidobacterium adolescentis persisted at higher pH values both in vitro and in vivo. Lactate and acetate were the main organic acids detected in both aspirates and fermentors. These data show that the simulator used in this investigation was capable of modeling the effects of environmental influences on the upper GI microbiota of EN patients and that gastric pH of <4 is not sufficient to prevent microbial overgrowth in these individuals.     

Bacterial translocation from the intestines

Bacterial translocation is defined as the passage of viable bacteria from the gastrointestinal (GI) tract through the mucosal epithelium to other sites, such as the mesenteric lymph nodes, spleen, liver and blood. This paper reviews results from animal models utilized to obtain information concerning the defense mechanisms operating in the healthy host to confine bacteria to the GI tract. Gnotobiotic and antibiotic-decontaminated mice colonized with particular bacteria demonstrated that the indigenous GI flora maintains an ecologic equilibrium to prevent intestinal bacterial overgrowth and translocation from the GI tract. Studies with athymic (nu/nu) mice, thymus-grafted (nu/nu) mice, neonatally thymectomized mice, and mice injected with immunosuppressive agents demonstrated that the host immune system is another defense mechanism inhibiting bacterial translocation from the GI tract. Ricinoleic acid given orally to mice disrupted the intestinal epithelial barrier allowing indigenous bacteria to translocate from the GI tract. Thus, bacterial translocation from the GI tract of healthy adult mice is inhibited by: (a) an intact intestinal epithelial barrier, (b) the host immune defense system, and (c) an indigenous GI flora maintaining ecological equilibrium to prevent bacterial overgrowth. Deficiencies in host defense mechanisms act synergistically to promote bacterial translocation from the GI tract as demonstrated by animal models with multiple alterations in host defenses. Bacterial translocation occurred to a greater degree in mice with streptozotocin-induced diabetes, mice receiving nonlethal thermal injury, and mice receiving the combination of an immunosuppressive agent plus an oral antibiotic than in mice with only a primary alteration in host defenses. The study of bacterial translocation in these complex models suggests that opportunistic infections from the GI tract occur in discrete stages. In the healthy adult animal, bacterial translocation from the GI tract either does not occur or occurs at a very low level and the host immune defenses eliminate the translocating bacteria. Bacterial translocation does take place if one of the host defense mechanisms is compromised, such as a deficiency in the immune response, bacterial overgrowth in the intestines, or an increase in the permeability of the intestinal barrier. In this first stage, the bacteria usually translocate in low numbers to the mesenteric lymph node, and sometimes spleen or liver, but do not multiply and spread systemically.(ABSTRACT TRUNCATED AT 400 WORDS)     

Acidomucin goblet cell expansion induced by parenteral nutrition in the small intestine of piglets

Total parenteral nutrition (TPN) impairs small intestine development and is associated with barrier failure, inflammation, and acidomucin goblet cell expansion in neonatal piglets. We examined the relationship between intestinal goblet cell expansion and molecular and cellular indices of inflammation in neonatal piglets receiving TPN, 80% parenteral + 20% enteral nutrition (PEN), or 100% enteral nutrition (control) for 3 or 7 days. Epithelial permeability, T cell numbers, TNF-alpha and IFN-gamma mRNA expression, and epithelial proliferation and apoptosis were compared with goblet cell numbers over time. Epithelial permeability was similar to control in the TPN and PEN jejunum at day 3 but increased in the TPN jejunum by day 7. By day 3, intestinal T cell numbers were increased in TPN but not in PEN piglets. However, goblet cell expansion was established by day 3 in both the TPN and PEN ileum. Neither TNF-alpha nor IFN-gamma mRNA expression in the TPN and PEN ileum correlated with goblet cell expansion. Thus goblet cell expansion occurred independently of overt inflammation but in association with parenteral feeding. These data support the hypothesis that goblet cell expansion represents an initial defense triggered by reduced epithelial renewal to prevent intestinal barrier failure.     

Effect of pH on an In Vitro Model of Gastric Microbiota in Enteral Nutrition Patients

Patients with dysphagia due to oropharyngeal disease or cerebrovascular accident require long-term nutritional support via enteral feeding, which often results in microbial overgrowth in the upper gastrointestinal (GI) tract. Gastric acid is the primary innate defense mechanism in the stomach and has been assumed to provide an effective barrier to microbial colonization at pH values of <4. To evaluate the efficacy of gastric acid as a barrier to overgrowth, the microbiota of gastric and duodenal aspirates was assessed by culturing methods. Additionally, a fermentor-based model incorporating enteral nutrition tubing of the gastric microbiota of enteral nutrition (EN) patients was constructed to assess the effect of pH on the microbiota. Results showed that gastric acidity had a relatively small effect on the numbers of microorganisms recovered from intestinal aspirates but did influence microbiota composition. Similarly, at pH 3 in the fermentor, a complex microbiota developed in the planktonic phase and in biofilms. The effect of pH on microbiota composition was similar in aspirates and in the fermentors. Candidas and lactobacilli were aciduric, while recoveries of Escherichia coli and Klebsiella pneumoniae decreased as pH was reduced, although both were still present in significant numbers at pH 3. Only Staphylococcus aureus and Bifidobacterium adolescentis persisted at higher pH values both in vitro and in vivo. Lactate and acetate were the main organic acids detected in both aspirates and fermentors. These data show that the simulator used in this investigation was capable of modeling the effects of environmental influences on the upper GI microbiota of EN patients and that gastric pH of <4 is not sufficient to prevent microbial overgrowth in these individuals.     

Qualitative and quantitative comparison of gut bacterial colonization in enterally and parenterally fed neonatal pigs

Total parenteral nutrition (TPN) has been associated with mucosal atrophy, impaired gut barrier function, and translocation of luminal bacteria with resultant sepsis in preterm human infants. Currently, we examined the effects of enteral (ENT) or TPN treatments on translocation events in neonatal pigs and on colonization and composition of microbiota in the neonatal gut. Newborn, colostrum-deprived pigs (<24 hours old) were fitted with intravenous catheters and were fed either ENT (n = 13) or TPN (n = 13) for 7 days. After 7 days of treatment, pigs were euthanized and samples were collected for bacterial culture from the blood, intestinal tract and organs. ENT pigs had increased numbers of bacterial genera isolated, higher concentrations of bacteria (CFU/g), and increased colonization of all segments of the intestinal tract compared to the TPN pigs. Translocation of bacteria from the intestinal tract to tissues or blood was similar (8 of 13) for both groups. The ENT group had 1/13 positive for Clostridium difficile toxin A whereas the TPN group had 5/13. We concluded that ENT favored increased bacterial concentrations comprised of more speciation in the gastrointestinal tract compared to TPN, and that TPN-treated piglets were at higher risk of colonization by toxin-expressing strains of C. difficile.     

Enteral nutrition and hepatosplanchnic region in critically ill patients - friends or foes?

Enteral nutrition (EN) is a preferred way of feeding in critically ill patients unless obvious contraindications such as ileus or active gastrointestinal bleeding are present. Early enteral nutrition as compared to delayed EN or total parenteral nutrition decreases morbidity in postsurgical and trauma patients. The hepatosplanchnic region plays a pivotal role in the pathophysiology of sepsis and multiple organ dysfunction syndrome. The beneficial effects of EN on splanchnic perfusion and energy metabolism have been documented both in healthy volunteers and animal models of sepsis, hemorrhagic shock and burns. By contrast, EN may increase splanchnic metabolic demands, which in turn may lead to oxygen and/or energy demand/supply mismatch, especially when hyperemic response to EN is not preserved. Therefore, the timing of initiation and the dose of EN in patients with circulatory failure requiring vasoactive drugs are a matter of controversy. Interestingly, the results of recent clinical studies suggest that early enteral nutrition may not be harmful even in patients with circulatory compromise. Nevertheless, possible onset of serious complications, the non-occlusive bowel necrosis in particular, have to be kept in mind. Unfortunately, there is only a limited number of clinically applicable monitoring tools for the effects of enteral nutrition in critically ill patients.     

双歧杆菌四联活菌片 对重症脑卒中患者肠道功能的影响

目的探讨双歧杆菌四联活菌片对重症脑卒中患者肠道菌群及肠黏膜屏障功能的影响。方法选择重庆市人民医院2017年1月至2017年11月收治的100例重症脑卒中患者,随机将入选患者分为治疗组和对照组各50例。治疗组在常规肠内营养(EN)治疗基础上联用双歧杆菌四联活菌片治疗,对照组给予常规EN治疗。比较两组患者治疗前后营养状况、肠道菌群数量和肠黏膜屏障功能。比较两组患者治疗后消化道并发症发生率。结果治疗前,两组患者营养状况、肠道菌群数量和肠黏膜屏障功能差异无统计学意义(P0.05)。治疗后,两组患者营养状况指标白蛋白(ALB)、血红蛋白(Hb)和三头肌肌围(MAMC)水平,肠道有益菌(双歧杆菌、乳杆菌和拟杆菌)数量均有所上升,肠黏膜屏障功能指标二胺氧化酶(DAO)、D-乳酸水平及肠道有害菌数量(小梭菌和肠球菌)均降低。与对照组相比,治疗组患者ALB、Hb和MAMC水平,肠道双歧杆菌、乳杆菌和拟杆菌数量均显著升高,DAO、D-乳酸水平和小梭菌、肠球菌数量均显著降低,差异有统计学意义(P0.05)。治疗组患者消化道并发症发生率均低于对照组(P0.05)。结论双歧杆菌四联活菌片能够显著改善脑卒中患者的营养状况,有效调节菌群失衡,改善肠黏膜屏障功能,降低并发症发生。     

基于免疫学的胃肠道微生态与胃癌相关研究进展

21世纪,随着人类微生物基因组计划和人类肠道元基因组计划的开展,科学家们越来越关注存在于人体百万亿计的微生物,尤其是机体中最为复杂的胃肠道微生物。同时,肠道黏膜免疫学也是近年来备受关注的研究方向。肠道不仅是消化吸收的代谢场所也是重要的免疫器官,肠黏膜含有丰富的淋巴细胞,它们与肠道微生物相互作用,参与机体的免疫防御、免疫平衡和免疫监视。胃肠道微生态平衡发生紊乱会影响机体免疫应答反应,进而引起疾病的发生发展。本文从免疫学的角度来论述胃肠道微生物在肿瘤尤其是胃癌的发生和治疗中所扮演的角色。     

Glutamine treatment decreases plasma and lymph cytotoxicity during sepsis in rats

Glutamine (Gln) is considered as a conditionally essential amino acid. Pharmacological supplementation of Gln helps to maintain the intestinal mucosal barrier, modulate cytokine production, and prevent organ injury during sepsis. Our previous study demonstrated the different effects of Gln on macrophage cytokine production in vitro or in vivo. The purpose of this study was to investigate the potential mechanism of Gln treatment to protect cells and modulate inflammation during sepsis in vivo. The results showed that administration of Gln significantly attenuated plasma-induced macrophage cytokine production and endothelial cell necrosis after cecal ligation and puncture in rats. In addition, it preserved human umbilical vein endothelial cell (HUVEC) viability and migration ability. Gln treatment also reduced lymph cytotoxicity by restoring macrophage tumor necrosis factor-α production, maintaining HUVEC viability, and decreasing endothelial cell necrosis. Mesenteric lymph duct ligation did not alleviate plasma cytotoxicity. Plasma lipopolysaccharide and d-lactate levels were suppressed after Gln treatment. Taken together, these results indicated that Gln administration can protect cells by attenuating the cytotoxicity of plasma and mesenteric lymph during sepsis.     

Probiotic research in neonates with congenital gastrointestinal surgical conditions – Now is the time

Neonates with congenital gastrointestinal surgical conditions (CGISC) receive parenteral nutrition, get exposed to multiple courses of antibiotics, undergo invasive procedures, and are nursed in intensive care units. They do not receive early enteral feeding and have limited opportunities for skin to skin contact with their mothers. Many of these infants receive gastric acid suppression therapies. All these factors increase the risk of gut dysbiosis in these infants. Gut dysbiosis is known to be associated with increased risk of infections and other morbidities in ICU patients. Experimental studies have shown that probiotics inhibit gut colonization with pathogenic bacteria, enhance gut barrier function, facilitate colonization with healthy commensals, protect from enteropathogenic infection through production of acetate, reduce antimicrobial resistance, enhance innate immunity, and increase the maturation of the enteric nervous system and promote gut peristalsis. Through these mechanisms, probiotics have the potential to decrease the risk of sepsis and inflammation, improve feed tolerance and minimise cholestasis in neonates with CGISC. Among preterm non-surgical infants, evidence from more than 35 RCTs and multiple observational studies have shown probiotics to be safe and beneficial. A RCT in neonates (N=24) with gastroschisis found that probiotic supplementation partially attenuated gut dysbiosis. Two ongoing RCTs (total N=168) in neonates with gastrointestinal surgical conditions are expected to provide feasibility data to enable the conduct of large RCTs. Rigorous quality assurance of the probiotic product, ongoing microbial surveillance and clinical vigilance are warranted while conducting such RCTs.     

肠黏膜屏障与肠道菌群的相互关系

摘要：人类肠道中微生物群与肠道环境相互作用以维持机体健康。肠黏膜屏障主要由黏液层、肠道菌群、肠道免疫系统和肠上皮细胞本身的完整性等构成。肠道作为直接与大量菌群接触的器官,其屏障功能在肠道健康中的作用尤为显著。肠道菌群与肠道屏障相互作用,保持肠道菌群与肠道屏障相对稳定,肠道菌群参与肠道免疫反应的建立,共同建立机体天然防御系统,在保持肠道免疫的动态平衡中具有重要作用。当两者之间的平衡被打破时,可诱发功能性胃肠病（如肠易激综合征）及免疫相关性疾病（如炎症性肠病）。本文主要阐述肠黏膜屏障与肠道菌群之间的相互关系以及与肠道屏障功能障碍相关的肠道疾病。     

Gastrointestinal absorption and metabolism of apple polyphenols ex vivo by the pig intestinal mucosa in the Ussing chamber

Polyphenols contained in food have various positive effects on human health. The absorption and metabolism of polyphenols in the intestinal tract needs to be studied to estimate these effects. The Ussing chamber technique was used to investigate the transport behavior of apple polyphenols through pig small intestinal mucosa, which served as a model for human gastrointestinal mucosa. The identities and concentrations of polyphenols and their metabolites in the half-chambers (luminal and basolateral) within an incubation period of 4 h were determined by HPLC–MS/MS and HPLC–DAD (DAD = diode-array detection). Flux values were also measured. It was found that 5-caffeoylquinic acid and caffeic acid were absorbed and translocated to the basolateral side (1.9 and 3.7%, respectively), but other compounds, including glycosides of phloretin and quercetin, were observed without translocation. A Ussing chamber utilizing pig small intestinal mucosa is a suitable model for assessing the effect of apple polyphenols on mucosal integrity and nutrition absorption across porcine mucosa.     

Ghrelin,CGRP,NT对胃肠作用的研究进展

胃肠道是人体内最大的激素分泌器官,是调节肽即胃肠激素最丰富的来源。胃肠激素与胃肠功能有很大关系,它们与神经系统一起,共同调节消化器官的运动、分泌和吸收及其他多种功能。促生长素(Ghrelin)、降钙素基因相关肽(CGRP)和神经降压素(NT)是近年来新发现的胃肠激素中的代表。Ghrelin主要由胃组织产生,可以促进胃肠蠕动,还可促进胃酸分泌,这些作用是由迷走神经所介导的,ghrelin还具有对消化道粘膜的保护作用,此作用受多种方式调控。CGRP广泛分布于中枢和外周神经系统,有调节胃肠血流、胃肠分泌及胃肠运动等多种功能,目前学者普遍认为CGRP这些生物学效应的发挥是通过一氧化氮(NO)及前列腺素(PG)介导的。NT广泛分布于脑和胃肠道及其它组织中,由肠道N细胞分泌,能够抑制胃肠运动,对胃肠黏膜细胞具有保护作用,这些作用是迷走神经、调节肽等多种途径介导的。随着对这三种胃肠激素的深入了解,人们将对人体胃肠道疾病产生更加深刻的认识。本文就近年来对Ghrelin、CGRP、NT对胃肠作用的研究作一综述。     

Effect of temperature on proteinase activities of enteral microbiota and intestinal mucosa of fish of different ecological group

Effect of temperature on proteinases activities of enteral microbiota and of intestinal mucosa was studied in five fish species (roach Rutilus rutilus, crucian carp Carassius carassius, common perch Perca fluviatilis, pike-perch Zander lucioperca, and pike Esox lucius) belonging by the nutrition type to different ecological groups. Essential differences of temperature characteristics of proteinases of intestinal mucosa and of enteral microbiota are revealed in fish belonging by the nutrition type to different ecologic groups. The character of the t0-function of proteinases of intestinal mucosa and enteral microbiota by casein and hemoglobin as a rule is different. The highest values of relative proteinases activities for casein in the zone of low temperatures (38 and 45.3 % of the maximal activity) are found at study of proteinases of enteral microbiota in common perch and crucian carp. The latter indicates a significant adaptability of the enteral microbiota proteinases of common perch and crucial carp to functioning at low temperatures.     

Erythropoietin protects intestinal epithelial barrier function and lowers the incidence of experimental neonatal necrotizing enterocolitis

The impermeant nature of the intestinal barrier is maintained by tight junctions (TJs) formed between adjacent intestinal epithelial cells. Disruption of TJs and loss of barrier function are associated with a number of gastrointestinal diseases, including neonatal necrotizing enterocolitis (NEC), the leading cause of death from gastrointestinal diseases in preterm infants. Human milk is protective against NEC, and the human milk factor erythropoietin (Epo) has been shown to protect endothelial cell-cell and blood-brain barriers. We hypothesized that Epo may also protect intestinal epithelial barriers, thereby lowering the incidence of NEC. Our data demonstrate that Epo protects enterocyte barrier function by supporting expression of the TJ protein ZO-1. As immaturity is a key factor in NEC, Epo regulation of ZO-1 in the human fetal immature H4 intestinal epithelial cell line was examined and demonstrated Epo-stimulated ZO-1 expression in a dose-dependent manner through the PI3K/Akt pathway. In a rat NEC model, oral administration of Epo lowered the incidence of NEC from 45 to 23% with statistical significance. In addition, Epo treatment protected intestinal barrier function and prevented loss of ZO-1 at the TJs in vivo. These effects were associated with elevated Akt phosphorylation in the intestine. This study reveals a novel role of Epo in the regulation of intestinal epithelial TJs and barrier function and suggests the possible use of enteral Epo as a therapeutic agent for gut diseases.     

OPC-compounds prevent oxidant-induced carbonylation and depolymerization of the F-actin cytoskeleton and intestinal barrier hyperpermeability

Rebamipide (OPC-12759), a quinolone derivative, and OPC-6535, a thiazol-carboxylic acid derivative, are compounds with ability to protect gastrointestinal (GI) mucosal integrity against reactive oxygen metabolites (ROM). The underlying mechanism of OPC-mediated protection remains poorly understood. It is now established that ROM can injure the mucosa by disruption of the cytoskeletal network, a key component of mucosal barrier integrity. We, therefore, investigated whether OPC compounds prevent the oxidation, disassembly, and instability of the cytoskeletal protein actin and, in turn, protect intestinal barrier function against ROM. Human intestinal (Caco-2) cell monolayers were pretreated with OPC (-12759 or -6535) prior to incubation with ROM (H2O2) or HOCl). Effects on cell integrity (ethidium homodimer-1), epithelial barrier function (fluorescein sulfonic acid clearance), and actin cytoskeletal integrity (high-resolution laser confocal) were then determined. Cells were also processed for quantitative immunoblotting of G- and F-actin to measure oxidation (carbonylation) and disassembly of actin. In monolayers exposed to ROM, preincubation with OPC compounds prevented actin oxidation, decreased depolymerized G-actin, and enhanced the stable F-actin. Concomitantly, OPC agents abolished both actin cytoskeletal disruption and monolayer barrier dysfunction. Data suggest for the first time that OPC drugs prevent oxidation of actin and lead to the protection of actin cytoskeleton and intestinal barrier integrity against oxidant insult. Accordingly, these compounds may be used as novel therapeutic agents for the treatment of a variety of oxidative inflammatory intestinal disorders with an abnormal mucosal barrier such as inflammatory bowel disease.     

Tissue distribution and plasma clearance of heparin-binding EGF-like growth factor (HB-EGF) in adult and newborn rats

Heparin-binding EGF-like growth factor (HB-EGF), a member of the epidermal growth factor (EGF) family, can protect intestinal epithelial cells from various forms of injury in vitro and attenuate intestinal ischemia/reperfusion damage in vivo. With the goal of eventual clinical use of HB-EGF to protect the intestines from injury in neonates, children, and adults, the pharmacokinetics and biodistribution of 125I-labeled HB-EGF were investigated. After intravenous bolus, HB-EGF had a distribution half-life of 0.8 min and an elimination half-life of 26.67 min. After gastric administration, the bioavailability was 7.8%, with a 2.38 h half-life in the absorption phase and an 11.13 h half-life in the elimination phase. After intravenous dosing, most radioactivity was found in the plasma, liver, kidneys, bile, and urine, whereas it was mainly distributed in the gastrointestinal tract after intragastric administration. The degradation of 125I-HB-EGF in plasma from newborn rats was lower than that in adult rats after gastric administration. This supports the feasibility of enteral administration of HB-EGF in the treatment of gastrointestinal diseases, including newborns afflicted with necrotizing enterocolitis.     

消化道肿瘤术后肠道菌群失调的诊治

目的 :探讨消化道肿瘤术后肠道菌群失调的诊断与治疗。方法 :收集 10 4例消化道肿瘤患者 ,并分别给以常规补液、肠内营养及肠外营养。结果 :17例患者术后发生腹泻 ,肠内营养组无术后肠道菌群失调(P<0 .0 5 ) ,常规补液组及肠外营养组肠道菌群失调症患者 14例 ,给以双歧杆菌制剂等治疗菌群失调 ,症状得以改善。结论 :肠内营养有防治肠道菌群失调的作用 ,双歧杆菌等微生态制剂可用于治疗消化道肿瘤术后肠道菌群失调。     

多糖对肠道功能调节作用的研究进展

肠道是机体重要的消化器官,亦是共生微生物群的主要寄居场所,在维持机体正常生命活动如免疫和内分泌功能中发挥着重要作用。 肠道功能紊乱与疾病的发生以及发展过程密切相关。近年来,多项研究结果显示,多糖具有肠道功能调节作用,包括通过作用于肠道黏膜 参与机体免疫过程、保护肠道屏障结构和功能的完整性、调节肠道菌群组成以及刺激肠道内分泌。从伴随疾病过程中的肠道功能紊乱的角度, 对多糖调节肠道功能的作用机制进行综述。     

大黄防治胃肠黏膜屏障损伤的研究进展

临床上许多疾病的发生、发展与预后都与胃肠黏膜屏障损伤有关,机体在应激状态下,胃肠黏膜屏障功能破坏,细菌、内毒素易位入血,形成肠源性感染,引起全身炎症反应(SIRS),最终引发多器官功能障碍综合症(MODS)。肠道黏膜屏障主要由四部分组成:即肠道黏膜的机械屏障、化学屏障、生物屏障和免疫屏障。中药大黄具有通里攻下、清热解毒、活血化瘀的功用,发挥着多环节、多靶点的作用。已有研究表明:大黄对胃肠黏膜屏障各层结构均有明显保护作用,本文现将胃肠黏膜屏障损伤发生机制和大黄防治机理作一综述。