Administration of nitric oxide with caspase inhibitors minimizes bacterial translocation in experimental intestinal transplantation

BACKGROUND: Bacterial translocation (BT) has been suggested to be responsible for the high incidence of infections occurring after small-bowel transplantation (Trp). Nitric oxide (NO) and apoptosis could affect cell demise. The aim of this study was to asses whether supplementation of University of Wisconsin (UW) solution with NO donors and apoptosis inhibitors can abolish BT in Trp. METHODS: The following experimental groups were studied: sham, Trp, intestinal transplantation, Trp+spermine NONOate (NONOs), and Trp+NONOs+caspase inhibitor Z-Val-Ala-Asp(Ome)-fluoromethylketone(Z-VAD-fmk). Histologic analysis, caspase-3 activity, DNA fragmentation, and BT from graft to mesenteric lymph nodes, liver, and spleen were measured in tissue samples after transplantation. RESULTS: During intestinal transplantation, apoptosis and necrosis were increased, showing graft injury and high levels of BT. The rats treated with NONOs showed a histologic protection of transplanted graft and a decrease in BT despite caspase-3 and DNA fragmentation-inducing effects. Administration of caspase inhibitor Z-VAD to NONOs-treated rats reversed the NO apoptosis-inducing effects and showed the lowest levels of BT in all tissues. CONCLUSIONS: Exogenous administration of NO associated with the inhibition of apoptosis maintains the graft in optimal conditions in terms of BT and improves the histology of the graft after intestinal transplantation in rats.     

C-reactive protein may be a marker of bacterial translocation in experimental intestinal obstruction

BACKGROUND: C-reactive protein (CRP) is used as a marker of intestinal ischaemia. This study evaluated whether CRP levels can be used to detect ischaemia-induced (strangulated) intestinal obstruction and subsequent bacterial translocation. METHODS: Forty-eight rats, divided into four groups underwent the following procedures: anaesthesia alone (native controls), laparotomy (sham-operated controls), or surgical induction of simple or strangulated intestinal obstruction (simple and strangulated obstruction groups, respectively). Blood samples were collected for culture and serum CRP analysis. In addition, liver and mesenteric lymph node (MLN) specimens were collected for culture, to determine the presence of bacterial translocation; and ileal segments, for histopathological investigation. RESULTS: CRP levels and rates of bacterial translocation, expressed as colony forming units (cfu) per gram wet tissue, were higher in both intestinal obstruction groups than in the native and sham-operated control groups (P < 0.001 for both). The increases in CRP levels paralleled increases in the number of cfu in the MLN and liver cultures (P < 0.01). Compared to controls, animals in the obstruction groups also had a higher incidence of positive blood cultures (P < 0.005) and greater histopathologic evidence of inflammatory infiltration of the lamina propria (P < 0.01). However, no significant difference between the simple and strangulated obstruction groups was observed. CONCLUSION: CRP levels increase with the severity of bacterial translocation in acute intestinal obstruction but do not permit discrimination between simple and strangulated intestinal obstruction.     

78例肝硬化患者肠道细菌易位及其相关性研究

目的了解肝硬化患者肠道细菌易位（BT）的发生率及其相关危险因素,分析BT与术后感染的关系。方法对77例肝移植和1例未行肝移植的肝硬化患者进行术中采样,取外周血、门脉血及肠系膜淋巴结（MLN）,分别进行需氧及厌氧培养,了解BT的发生率。结合术前、术后各种临床资料分析发生BT的危险因素及BT的临床意义。结果78例肝硬化患者中BT的发生率为10．3％（8／78）;细菌易位的部位以MLN为主,占5／8,发生BT的细菌主要是肠道G-兼性厌氧杆菌（55．6％）,其次为G^＋兼性厌氧球菌（22．2％）。BT组患者术前胆红素总量显著高于无BT组（P=0．022）;发生BT的患者其术后感染的风险是无BT患者的1．3倍。结论高胆红素血症是促发BT的独立危险因素,发生BT的肝移植患者术后感染的风险明显升高。     

Bacterial translocation in clinical intestinal transplantation

BACKGROUND: Bacterial translocation (BT) has been suggested to be responsible for the high incidence of infections occurring after small bowel transplantation (SBTx). Bacterial overgrowth, alteration of the mucosal barrier function as a consequence of preservation injury or acute rejection (AR), and potent immunosuppression are all associated with BT. The aim of this study was to evaluate and quantify the correlation of BT with these events. METHODS: Fifty pediatric SBTx recipients on tacrolimus and prednisone immunosuppression were analyzed. Blood, stool, and liver biopsies and peritoneal fluid were cultured (circa 4000 total specimens) when infection was clinically suspected or as part of follow-up. BT episodes were considered when microorganisms were found simultaneously in blood or liver biopsy and stool. RESULTS: BT (average of 2.0 episodes/patient) was evident in 44% of patients and was most frequently caused by Enterococcus, Staphylococcus, Enterobacter, and Klebsiella. The presence of a colon allograft was associated with a higher rate of BT (75% vs. 33.3%). Furthermore, the transplantation procedure (colon vs. no colon) affected the rate of BT: SBTx=40% vs. 25%, combined liver and SBTx=100% vs. 30%, multivisceral transplantation=25% vs. 50%. AR was associated with 39% of BT episodes. BT followed AR in 9.6% of the cases. In 5.2% of the cases, positive blood cultures without stool confirmation of the bacteria were seen. Prolonged cold ischemia time (CIT) affected BT rate significantly (CIT>9 hr 76% vs. CIT<9 hr 20.8%). CONCLUSIONS: This study shows that 1) a substantial percentage of, but not all, BT is associated with AR, 2) the presence of a colon allograft increases the risk for BT, and 3) a long CIT is associated with a high incidence of BT after SBTx.     

Simple intestinal obstruction causes bacterial translocation in man

Indirect clinical evidence has accumulated indicating that the gut may be a reservoir for microorganisms causing systemic infection in man. Our experimental results, in a variety of animal models, demonstrate that bacteria can translocate across the mucosal barrier and cause systemic infections. To determine directly whether bacterial translocation occurs in man, we cultured mesenteric lymph nodes (MLNs) obtained at laparotomy from 42 patients, none of whom were clinically infected. Ten (59%) of 17 patients with intestinal obstruction (none of whom had necrotic bowel) had bacteria in their MLNs, in contrast to one (4%) of 25 patients operated on for other reasons. The most common bacteria cultured from the MLNs was Escherichia coli. Thus, it appeared that simple intestinal obstruction of the colon or small bowel in the absence of necrotic bowel was associated with bacterial translocation.     

急性坏死性胰腺炎时肠通透性改变与细菌移位的实验研究

通过复制犬急性坏死性胰腺炎（ＡＮＰ）模型，探讨ＡＮＰ时肠通透性变化及其与肠道细菌．移位的关系。杂种犬１５只，分ＡＮＰ组（ｎ＝８）和对照组（ｎ＝７）。气相色谱法测定尿中乳果糖和甘露醇的含量；每日血培养；第７天活杀后各组脏器标本作细菌培养。结果发现，ＡＮＰ组尿中乳果糖／甘露醇比率高出对照组２～１２倍，发病后第２天最为显著（Ｐ＜０．０１），ＡＮＰ犬都出现了肠道细菌移位，血培养阳性率为８／８，以发病后４８小时内最高，镜检证实ＡＮＰ组肠粘膜上皮绒毛脱落。对照组只有２只犬在肠系膜淋巴结中培养出细菌，其它脏器和血培养全部阴性。本研究提示：ＡＮＰ后４８小时肠通透性即出现病理性升高，肠粘膜屏障遭到严重破坏，发生肠道细菌移位。     

Patterns of bacterial translocation in experimental biliary obstruction

INTRODUCTION: Biliary obstruction is associated with impaired intestinal barrier function and translocation of enteric bacteria to the systemic circulation. Traditional live culture techniques may overlook translocation of dead bacterial fragments that stimulate the inflammatory response. The aim of this study was to estimate the extent and pattern of bacterial translocation in experimental biliary obstruction. MATERIALS AND METHODS: Thirty 9-week-old male Wistar rats were randomized to undergo bile duct ligation (BDL, n = 20) or sham operation (n = 10). Seven days after operation, each animal received 1 ml of (111)indium-oxyquinolone-labeled Escherichia coli p.o. Samples of liver, spleen, mesenteric lymph nodes, and lung were harvested 4 h later and analyzed for live bacteria and (111)indium activity. RESULTS: There was significantly more live bacterial translocation detected in BDL animals than in sham-operated animals (P = 0.00008, chi(2)). Labeled bacterial fragments were detected in all locations sampled in all animals. Sham-operated animals had significantly more labeled bacterial fragments detected in the liver (P = 0.0001) and the spleen (P = 0.03) than the BDL animals. The mean total bacterial survival in the BDL group was 30 +/- 13% and 0% in the sham operated group. CONCLUSION: These results demonstrate that non-viable bacterial fragments are present in sterile extra-intestinal sites in normal animals and that translocation of live bacteria is markedly increased in experimental biliary obstruction. These results also suggest that failure of bacterial killing is an important factor facilitating bacterial translocation in the presence of established biliary obstruction.     

Angiotensin II plays a role in acute murine experimental autoimmune cystitis

OBJECTIVES: To investigate whether angiotensin II (AII) receptor antagonism decreases the inflammation and oedema in acute murine experimental autoimmune cystitis (EAC), as interstitial cystitis (IC) might have an autoimmune component and AII has been implicated in autoimmune-mediated vascular congestion, oedema and scarring. MATERIALS AND METHODS: Female Balb/cAN mice were divided into three treatment groups (eight in each group) that were autoimmunized with bladder homogenate to induce EAC. One group received an AII type 1 receptor (AT(1)) antagonist, one group an AII type 2 receptor (AT(2)) antagonist, and one group remained untreated (EAC). A control and sham-injected group were also included. After 10 weeks, bladders were removed, sectioned, and stained with haematoxylin and eosin. RESULTS: Grossly, there was no thickening or adhesions in the bladders of the control or sham-injected mice. In five of seven surviving EAC bladders, there were dense adhesions to surrounding peritoneal structures. There were also adhesions and bladder thickening in all of the AT(2) antagonist-treated mice (though in a milder form) but in only two of seven surviving AT(1) antagonist-treated mice. There was no inflammation or oedema in the sham and control groups. All the EAC bladders were inflamed, with submucosal oedema and urothelial detachment from the lamina propria. In the AT(1) antagonist-treated mice there was no inflammation or oedema. By contrast, all AT(2) antagonist-treated mice had moderate inflammation and minor detachment of the urothelium from the lamina propria. CONCLUSIONS: AT(1) receptor blockade ameliorated the inflammatory infiltration, submucosal oedema, and urothelial detachment associated with EAC in mice. This was achieved to a lesser extent by AT(2) receptor blockade. If some patients with IC have a pathophysiology similar to that of EAC mice, there might be potential benefit from AII receptor blockade.     

Treatment with Met-RANTES decreases bacterial translocation in experimental colitis

BACKGROUND: During colitis, epithelial function is impaired, leading to increased bacterial translocation. Recent studies have shown the important role of proinflammatory cytokines and chemokines, including RANTES (regulated on activation, normal T-cell expressed and secreted), in inflammatory bowel diseases (IBDs). In this study, we evaluated the role of Met-RANTES, an antagonist of the RANTES receptor, on the impairment of bacterial translocation in a rat model of colitis. METHODS: Rats were randomly assigned to 3 groups. Group 1 = control, group 2 = experimental colitis, and group 3 = colitis plus Met-RANTES treatment. On day 7 after colitis was induced, plasma tumor necrosis factor-alpha colon tissue myeloperoxidase and portal blood endotoxin levels were measured. Lymph node, liver, and spleen culture quantified bacterial translocation. RESULTS: Met-RANTES treatment resulted in significant decreases in colonic damage as well as bacterial translocation in experimental colitis. CONCLUSIONS: These results suggest that chemokine receptor antagonists may potentially be useful in the treatment of IBDs.     

谷氨酰胺治疗肠梗阻的疗效分析

目的：谷氨酰胺（Gln）对肠梗阻肠黏膜通透性及细菌移位的影响。方法：将80例肠梗阻患者随机分为对照组和治疗组。对照组给予禁食、胃肠减压、纠正水电解质和酸碱平衡紊乱,全胃肠外营养以及抗生素等常规治疗;治疗组在常规治疗基础上,全胃肠外营养液中加用Gln。比较两组患者入院时、治疗5 d、治疗10 d的常规指标（体质量、血糖、血清白蛋白、血清前白蛋白、血清转铁蛋白）、免疫学指标（CRP、外周血白细胞计数I、gAI、gMI、gG、TNF-ɑI、L-6）和肠黏膜通透性评价指标[外周血浆Gln浓度、尿乳果糖甘露醇比值（L/M）、外周血细菌DNA阳性率]。结果：治疗后两组血清白蛋白、血清前白蛋白、血清转铁蛋白显著升高（P〈0.05）;血糖明显降低（P〈0.05）。对照组治疗10 d IgA较治疗5 d明显降低（P〈0.05）;治疗组无明显变化。治疗后治疗组血浆Gln浓度高于对照组（P〈0.01）,治疗组治疗10 d尿L/M、外周血细菌DNA阳性率低于对照组（P〈0.01）。结论：肠梗阻患者应用Gln有助于维护肠黏膜屏障功能、减少细菌移位。     

Effects of quercitrin on bacterial translocation in a rat model of experimental colitis

Background

This study aimed to analyze the effects of quercitrin, which has anti-inflammatory properties, on bacterial translocation in inflammatory bowel diseases by using an experimental colitis model.

大鼠急性胰腺炎肠道细菌易位的实验研究

目的本实验是观察大鼠急性胰腺炎后经胃肠道给予庆大霉素或谷氨酰胺对肠道细菌易位的影响。方法结扎封闭群 Wistar 大鼠的胆管,制成急性胰腺炎模型。分假手术组、急性胰腺炎组、庆大霉素组和谷氨酰胺组4组,各组又分5个亚组,分别于术后24、48、72、96和144小时处死动物。无菌下取回盲部淋巴结、胰腺、脾、肝、门静脉血和盲肠内容做细菌培养、计数。胰腺和小肠组织做病理检查。结果表明大鼠急性胰腺炎发生后,肠系膜淋巴结细菌数和阳性率明显增高,与门静脉血比较有显著差异。庆大霉索组盲肠内容和肠系膜淋巴结的大肠杆菌数明显减少,革兰氏阳性菌明显增多。谷氨酰胺组盲肠内容和肠系膜淋巴结的细菌数均显著减少。结论大鼠急性胰腺炎发生后早期肠道细菌易位主要经肠系膜淋巴途径。经胃肠道给予谷氨酰胺可防止肠道内细菌易位,减少胰腺感染的发生。     

Effects of somatostatin analogues and vitamin C on bacterial translocation in an experimental intestinal obstruction model of rats.

The passage of viable endogenous bacteria and their products across the intact intestinal mucosal barrier, disseminating to the mesenteric lymph nodes, peritoneal cavity, spleen, liver, and circulation, is defined as bacterial translocation. Intestinal obstruction induces bacterial translocation due to mucosal disruption, motility dysfunction, and increased intestinal volume, leading to bacterial overgrowth. In a rat model of intestinal obstruction, the effects of both high-dose vitamin C (350 microg/kg), an antioxidant agent known to have a cytoprotective effect in ischemia-reperfusion injury, and somatostatin (20 microg/kg), a gastrointestinal antisecretory agent, in preventing bacterial translocation were studied. Both intestinal and liver samples from the rats was observed, and it was found that the rate of bacterial translocation was 100% in the control group, and only 43% for the rats who were given intraperitoneal vitamin C and somatostatin. The difference was statistically significant. In conclusion, we are convinced that vitamin C and somatostatin analogues may have protective effects against bacterial translocation in mechanical bowel obstruction.     

Prostaglandin E1 maintains structural integrity of intestinal mucosa and prevents bacterial translocation during experimental obstructive jaundice.

The absence of bile in the gut lumen induces mucosal injury and promotes bacterial translocation (BT). Prostaglandin E (PGE) has a protective effect on the mucosal layer of the alimentary tract. We hypothesize that PGE1 may prevent BT by its beneficial action on the mucosa of the small bowel. Thirty Wistar albino rats were divided equally into 3 groups; Group 1 (control) underwent sham laparotomy, group 2 obstructive jaundice (OJ) and group 3 (OJ + PGE1) underwent common bile duct (CBD) ligation and transection. Groups 1 and 2 received; 1 mL normal saline and group 3 received 40 mg of the PGE1 analogue misoprostol dissolved in 1 mL normal saline administered by orogastric tube once daily. After 7 days, laparotomy and collection of samples for laboratory analyses were performed, including bacteriological analysis of intestine, mesenteric lymph nodes (MLNs), and blood, and histopathologic examination of intestinal mucosa to determine mucosal thickness and structural damage. Serum bilirubin and alkaline phosphatase levels confirmed OJ in all animals with CBD transection. The mucosal damage score was significantly reduced in jaundiced animals receiving PGE1 compared to jaundiced controls (2.15 +/- 0.74 vs 5.3 +/- 0.59; p < .00001) and mucosal thickness was greater (607 +/- 59.1 microm vs. 393 +/- 40.3 microm; p < .00001). The incidence of BT to MLNs decreased from 90% to 30% (p < .02) when jaundiced rats received PGE1. PGE1 treatment reduced the detection rate of viable enteric bacteria in the blood from 60% to 10% (p < .057). We conclude that administration of PGE1 provides protection against OJ-induced atrophy and damage of intestinal mucosa, and thereby prevents translocation of enteric bacteria to underlying tissues.     

Role of parenteral nutrition in preventing malnutrition and decreasing bacterial translocation to liver in obstructive jaundice

Surgery in patients with obstructive jaundice is associated with significant infectious complications probably due to impaired immune function and malnutrition. Total parenteral nutrition (TPN) may alleviate malnutrition but may also promote bacterial translocation (BT) from the gut. To elucidate if TPN can prevent malnutrition without promotion of BT in obstructive jaundice, 40 dogs underwent laparotomy for tissue sampling and placement of a central venous line and were allocated into one of four groups: I (PO-control) received dog chow and water ad libitum; II (PO-CBDL) underwent ligation of common bile duct (CBDL) and was fed dog chow; III (TPN-control) received TPN; and IV (TPN-CBDL) underwent CBDL and received TPN. Body weight, blood samples for liver function tests and bacterial culture, and tissues from liver and mesenteric lymph nodes (MLN) for quantitative bacterial culture and for histology were obtained prior to and 2 weeks after the experiment. The incidence of BT to MLN was 40% in the PO-CBDL and TPN-CBDL animals, which was significantly different from the other two groups (0%);p<0.05). The incidence of BT to liver was 70% (7/10) in the PO-CBDL animals, which was significantly higher than that in groups I, III, and IV (0%, 20%, 20%, respectively) (p<0.05). The PO-CBDL animals showed a significant decrease in body weight and prealbumin compatible with malnutrition, whereas the TPN-CBDL animals showed a significant increase in alkaline phosphatase and a consistent cholestasis on histology. The data suggest that TPN can prevent jaundice-associated malnutrition and decrease BT to liver but should be administered cautiously because it may precipitate cholestasis.

---

Resumen La cirugía en pacientes con ictericia obstructiva se asocia, en forma significativa, con complicaciones infecciosas, probablemente debido a alteración de la función inmune y a malnutrición. La nutrición parenteral total (NPT) puede mejorar la malnutricón, pero también puede promover la translocación bacteriana (TB) a partir del intestino. Con el objeto de dilucidar si la TPN es capaz de prevenir la malnutrición sin que promueva la TB en pacientes con ictericia obstructiva, realizamos laparotomía en 40 perros para obtener muestras de tejido y colocar una línea venosa central; los animales fueron asignados a uno de cuatro grupos: I (PO-control), el cual recibió comida canina y agua sin limitación; II (PO-LCC), en el cual se practicó ligadura del canal colédoco y recibió comida canina; III (PON-PT control), el cual recibió NPT; y IV (NPT-LCC), en el cual se practicó ligadura del canal colédoco y recibió NPT. Se determinó el peso corporal, se tomaron muestras sanguíneas para pruebas de función hepática y para cultivos bacteriológicos y muestras tisulares del hígado y de los ganglios linfáticos mesentéricos (GLM), para cultivos bacteriológicos cuantitativos y para histología antes y dos semanas después del experimento. La incidencia de TB a los GLM fue de 40% en los animales de los grupos PO-LCC y NPT-LCC, tasa significativamente diferente de la de los otros dos grupos (0%, p<0.05). La incidencia de TB al hígado fue de 70% (7/10) en los animales del grupo PO-LCC, significativamente más alta que la de los de los grupos I, III y IV (0%, 20%, 20%, respectivamente) (p<0.05). Los animales PO-LCC mostraron una disminución significativa en el peso corporal y en los niveles de prealbúmina, compatibles con malnutrition, en tanto que los animales NPTLCC exhibieron un significativo aumento en la fosfatasa alcalina y colestasis consistente en el examen histológico. Tales datos sugieren que la NPT puede prevenir la malnutrición asociada con malnutrición y disminuir la TB hacia el hígado, pero que debe ser cautelosamente administrado debido a que precipita colestasis.

---

Résumé La chirurgie chez le patient ayant un ictère par obstruction est associée à une morbidité infectieuse élevée, probablement en raison d'une détérioration des fonctions immunitaires et de la malnutrition. L'alimentation parentérale totale (APT) peut améliorer l'état nutritionnel mais elle peut également être responsable de translocation bactérienne (TB) à partir du tube digestif. Pour savoir si l'APT pouvait améliorer l'état nutritionnel sans promouvoir la TB, 40 chiens ont été laparotomisés pour placer une voie veineuse centrale, faire certains prélèvements tissulaires et ensuite, ont eu: 1) rien d'autre qu'une alimentation adaptée ad lib (goupe témoin: PO), 2) une ligature de la voie biliaire principale (groupe CBDL) et une alimentation adaptée, 3) une APT seule (groupe TPN-contrôle), et 4) une ligature de la voie biliaire principale suivie d'une APT (goupe TPN-CBDL). Pour chaque animal, on a déterminé le poids, la fonction hépatique et on a réalisé des hémocultures, une biopsie hépatique et un prélèvement d'un ganglion mésentérique (MLN) pour examen bactériologique et histologique, au moment de l'intervention et deux semaines après. L'incidence de TB au niveau d'un MLN a été de 40% dans les groupe PO-CBDL et TPN-CBDL, significatement différente de celles des deux autres groupes où l'incidence était nulle (p<0.05). L'incidence de TB au niveau du foie a été de 70% (7/10) chez les animaux PO-CBDL, significativement différente (p<0.05) de celle des groupes PO (0%), TPN-contrôle (20%) et TPN-CBDL (20%). Les animaux PO-CBDL ont perdu plus de poids et leur taux de préalbumine était plus bas, alors les animaux TPN-contrôle avaient une augmentation significative des phosphatases alcalines et une cholestase constante en histologie. Ces données suggèrent que l'APT peut prévenir la malnutrition due à l'ictère et diminuer la TB au foie, mais elle doit être administrée avec précaution en raison de son influence sur la choléstase.

     

The role of intestinal barrier failure and bacterial translocation in the development of systemic infection and multiple organ failure

Traditionally, evaluation of intestinal function has been limited largely to monitoring gastric pH and intestinal motility. This clinical approach has led clinicians to equate normal intestinal motility with normal intestinal function and to assume that if stress-induced gastric bleeding can be prevented, all will be well. However, it is becoming increasingly clear that the gastrointestinal tract is not a passive organ and that intestinal dysfunction is not limited to ileus and upper gastrointestinal bleeding. Instead, the gastrointestinal tract is recognized as having important endocrine, metabolic, immunologic, and barrier functions, as well as its traditional role in nutrient absorption. Over the last 5 years, there has been a resurgence of interest in the role of intestinal barrier failure in the development of systemic infection and multiple organ failure in the critically ill or injured patient.