Manipulation of nitric oxide in an animal model of acute liver injury. The impact on liver and intestinal function

Background: Nitric oxide may have a protective effect on the liver during endotoxemia and chronic inflammation. There is evidence that it maintains liver and intestinal tissue integrity during inflammatory processes. We evaluated the impact of altering nitric oxide release on acute liver injury; the associated gut injury and bacterial translocation; at different time intervals. Methods: An acute rat liver injury model induced by D-galactosamine was used. Sprague Dawley rats were divided into four main groups: normal control; acute liver injury control; acute liver injury + N-nitro-L-arginine methyl ester (L-NAME); acute liver injury + L-NAME + L-arginine. Each group was divided into three subgroups according to the different time intervals (6; 12; 24 hours) after the induction of the liver injury. Liver enzymes and bilirubin were evaluated; as well as bacterial translocation; cecal and colonic microflora; and histological study of liver; ileum and cecum. Results:Liver enzymes increased significantly at all time intervals in acute liver injury + L-NAME compared to liver injury control groups. Bacterial translocation increased significantly in liver injury + L-NAME groups; at 6 hours to the liver; at 12 hours to the liver and mesenteric lymph nodes (MLNs); and at 24 hours to arterial and portal blood; liver and MLNS. Inhibition of nitric oxide increased significantly the Enterobacteriaceae count in cecum compared to normal and liver injury control groups. The G-negative anaerobes increased significantly in the colon compared to the liver injury control group. Conclusion: Inhibition of nitric oxide in an acute liver injury model potentiates the liver injury as evidenced by increased appearance of hepatocellular necrosis and elevated liver enzymes and bilirubin. It increases the Enterobacteriaceae in both cecum and colon and G-negative anaerobes in the colon. It also increases bacterial translocation to extra-intestinal sites. The increased bacterial translocation could be one of the mechanisms potentiating liver injury and nitric oxide may be pathophysiologically involved. Further studies are required to confirm this hypothesis

Effect of Lactobacillus supplementation with and without arginine on liver damage and bacterial translocation in an acute liver injury model in the rat.

In acute liver failure following hepatitis, toxic insults, or after major liver surgery, there is an increased bacterial translocation from the gut. This may explain some of the infectious complications seen in these conditions. To elucidate mechanisms and find possible preventive measures, we investigated the effect of rectal administration of arginine and probiotic bacteria (Lactobacillus spp.) on bacterial translocation and the extent of liver failure. Sprague-Dawley rats were used and five different Lactobacillus strains (Lb. reuteri R2LC, Lb. rhamnosus DSM 6594 (= strain 271), Lb. plantarum DSM 9843 (= strain 299v), Lb. fermentum 8704:3 (= strain 245), and Lb. reuteri (= strain 108) were administered rectally once daily for 8 days with and without 2% arginine. Acute liver injury (ALI) was induced on the eighth day by intraperitoneal injection of D-galactosamine (1.1 g/kg body weight), and samples were collected after 24 and 48 hours. Bacterial translocation was evaluated by bacterial culture from portal and arterial blood, mesenteric lymph nodes, and liver tissue. Liver enzymes and bilirubin were evaluated in the serum. The bacterial load in the cecum and colon was determined and the liver histopathological changes were studied. There was no mortality at any time. The liver enzymes and bilirubin decreased in some of the groups supplemented with lactobacilli with and without arginine compared with the ALI control group. The incidence of bacterial translocation and the number of the translocated bacteria decreased significantly in some of the supplemented groups. Lb. plantarum + arginine administration significantly reduced the level of the released liver enzymes, hepatocellular necrosis and inflammatory cell infiltration, bacterial translocation, and the number of Enterobacteriaceae in the cecum and colon. Rectal administration of different Lactobacillus strains with and without arginine in an ALI model significantly modulates the extent of the liver failure and reduces bacterial translocation. Lb. plantarum DSM 9843 (= strain 299v) with or without arginine seemed superior to the other Lactobacillus strains. The beneficial effect of arginine administration alone indicates a possible role of nitric oxide and polyamines in this process, and the lactobacilli may execute their action via the same mechanisms or via bacterial antagonism and/or enhancement of systemic and intestinal mucosal immunity.

Effect of oral supplementation of lactobacilli on bacterial translocation in acute liver injury induced by D-galactosamine.

BACKGROUND/AIM: Bacterial infections and sepsis are frequent complications of acute liver injury, with a high share in the mortality and morbidity of this condition. Bacterial translocation from the gut may play an important role in the high rate of infections observed. In this experiment the effect of different oral supplementation on bacterial translocation was evaluated in acute liver injury induced by D-galactosamine in the rat. METHODS: Rats were given oral supplements of lactulose, neomycin, Lactobacillus reuteri R2LC, and Lactobacillus plantarum DSM 9843 for 1 week. Liver injury was induced by intraperitoneal administration of 1.1 g/kg D-galactosamine. Twenty-four hours later, rats were sacrificed and liver enzymes and histology, intestinal bacterial count and microflora, intestinal mucosal histology, DNA and RNA content, bacterial translocation to blood, mesenteric lymph nodes, and liver, and serum endotoxin were studied or measured. RESULTS: Lactulose was highly effective in prevention of liver injury and bacterial translocation. Neomycin and Lactobacillus plantarum DSM 9843 showed a moderate effect in prevention of liver injury and bacterial translocation. Intestinal bacterial count and microflora were affected by different treatment modalities. No endotoxin concentration was found in any of the experimental groups. Both lactobacilli could significantly improve the mucosal proliferative state. CONCLUSIONS: Oral supplementation of lactulose with anti-endotoxin effect could successfully prevent the liver injury and the subsequent bacterial translocation in acute liver injury induced by administration of D-galactosamine in the rat. This effect was irrespective of the intestinal bacterial alteration or mucosal proliferative state.

The effect of pretreatment with endotoxin and lactobacillus on bacterial translocation in acute liver injury.

OBJECTIVE: To evaluate the effect of pretreatment with endotoxin or Lactobacillus reuteri pretreatment on bacterial translocation after acute liver injury. DESIGN: Experimental study. SETTING: University department, Sweden. SUBJECTS: 96 Sprague-Dawley rats were divided into four groups of 24 each. Three of them received intraperitoneal D-galactosamine, the fourth received pyrogen free water and was used as the normal control. Twenty-four hours later the study was terminated and samples collected. INTERVENTIONS: Endotoxin and L reuteri R2LC were injected intraperitoneally three days, one week, and two weeks before induction of liver injury. MAIN OUTCOME MEASURES: Extent of liver injury and bacterial translocation to mesenteric lymph nodes, liver, and systemic blood. RESULTS: The extent of liver injury and rate of bacterial translocation were lower three days after pretreatment with endotoxin, than after pretreatment with L reuteri. There was no other difference among the other groups. High concentrations of serum endotoxin were detected three days after pretreatment with endotoxin. There were no significant changes in small intestinal and caecal bacterial counts. CONCLUSION: Pretreatment with endotoxin effectively prevented liver injury by D-galactosamine and subsequent bacterial translocation. Pretreatment with L reuteri had no beneficial effect.

Oral arginine supplementation in acute liver injury.

Acute liver failure is accompanied by a high rate of bacterial and septic complications. Arginine has a potent effect on the immune system and modulates bacterial clearance in septic models. We studied the effect of oral arginine supplementation on the extent of liver injury and the associated bacterial translocation in an acute liver injury model in rats. Sprague-Dawley rats were divided into normal, liver injury, and arginine supplemented groups. In the arginine group, 2% arginine was supplemented daily through a nasogastric tube for 8 d. Acute liver injury was induced on the eighth day by intraperitoneal injection of D-galactosamine (1.1 g/kg body wt). Samples were collected 24 h after the liver injury. In the arginine-supplemented group, alkaline phosphatase, bilirubin, and aspartate aminotransferase were reduced significantly compared with the acute liver injury control group. The results of bacterial translocation in the arginine-supplemented group showed a significantly reduced number of translocated bacteria to the liver and mesenteric lymph nodes than occurred in the acute liver injury group. The histological study of the liver in arginine-supplemented group showed scattered areas of hepatocellular necrosis and inflammatory cell infiltration, and in the acute liver injury group there were more and widespread hepatocellular necrosis and inflammatory cell infiltration. Oral supplementation of arginine in an acute liver injury model improves significantly the state of the liver injury and reduces bacterial translocation to the liver and mesenteric lymph nodes.

The effect of endotoxin and Lactobacillus pretreatment on peritoneal macrophage behavior in acute liver injury in rat.

The effect of endotoxin and Lactobacillus R2LC pretreatment in acute liver injury, induced by D-galactosamine was studied. Administration of D-galactosamine resulted in much higher luminescent activity than the control (49.24 mV vs 19.75 mV), while there was no change detected on the metabolic response to phorbol-12-myristate-13-acetate (PMA) stimulation in the calorimeter. Pretreatment of animals with intraperitoneal administration of heat-killed Lactobacillus resulted in higher oxygen free radical production (105.46 mV) and a much higher metabolic response to PMA in microcalorimeter (24.63 pW per cell vs 13.03 in the control). On the other hand pretreatment with endotoxin resulted in higher luminometric response to PMA compared to liver injury group (65.87 mV), and no response in the microcalorimeter at all. Phagocytic activity was decreased from 86 to 59% after administration of D-galactosamine, and was not significantly influenced by any of the pretreatment modalities. This experiment showed that pretreatment with Lactobacillus resulted in increased macrophage metabolic activity, while endotoxin pretreatment rendered the macrophages unresponsive to subsequent stimulation. These findings may explain why pretreatment with endotoxin is beneficial in acute liver injury induced by D-galactosamine, but not with gram-positive bacteria including Lactobacillus.

Dynamics of bacterial translocation in acute liver injury induced by D-galactosamine in rat.

Bacterial translocation may play a role in acute liver injuries as high rates of infectious and septic complications are observed in these clinical conditions. Increased passage of endotoxin and translocating bacteria not only potentiates the extent of liver injury, but may also play a determining role in its final outcome. In this paper the incidence of bacteria] translocation in acute liver injury in rats is evaluated with other important pathological changes observed at different time points after liver injury induced by intraperitoneal administration of D-galactosamine. The bacterial translocation to the blood and other extraintestinal sites starts 3 h after induction of liver injury and is not found to be related to light microscopic changes in the liver or ileal or cecal mucosa, detectable levels of endotoxin in the portal blood, or DNA changes in the small and large intestinal mucosa, but corresponds to the release of liver enzymes in the serum.

Bacterial translocation in acute liver injury induced by D-galactosamine.

Acute liver injury is associated with a high rate of infectious and septic complications. Most of these infections are produced by gram negative enteric bacteria. We evaluated bacterial translocation, intestinal permeability, blood flow, portal pressure, and intestinal microflora after induction of liver injury and 70% liver resection in the rat. The rate of translocation to both portal and arterial blood was 100% at 24 hours and 50% at 48 hours after liver resection compared with 83% to portal vein and 50% to aortic blood at both time points after acute liver injury. Translocation to intraabdominal organs (liver, spleen, and mesenteric lymph nodes) was 100% in both groups at both 24 and 48 hours. The rate of translocation increased after liver injury at 48 hours with progression of the liver injury but was decreased in the 70% liver resection group with improvement of liver function. "Total aerobic" and "total anaerobic" bacterial counts in small intestine and cecum were not affected. Pulmonary, distal small intestine, and cecal blood flow were decreased in both groups, whereas blood flow in the proximal small intestine was unaffected. Portal pressure and flow were increased after 70% liver resection, but they were decreased in acute liver injury. After acute liver injury, permeability of both distal small intestine and cecum increased, but after liver resection only cecal permeability increased. The results of this experiment show that bacterial translocation occurs in experimental acute liver injury and that its dynamic, pattern and fate are different from that observed after liver resection, which is a reversible surgical model of liver insufficiency.

Reticuloendothelial system function in acute liver injury induced by D-galactosamine.

AIMS/METHODS: Reticuloendothelial system function, as assessed by clearance of radiolabelled bacteria, was evaluated in acute liver injury induced by D-galactosamine in rats, and compared with that after 70% liver resection model. RESULTS: Reticuloendothelial system function was significantly impaired in both instances, but the extent and the pattern of reticuloendothelial system impairment differed in the two models. While the elimination rate of the radiolabelled bacteria (k-value) decreased in both the liver resection and D-galactosamine groups (19% and 52%, respectively), the corrected phagocytic index (alpha) increased in 70% liver resection (247%), indicatine increased activity among the remaining reticuloendothelial system cells of the liver. Estimation of subserosal organ blood flow showed decreased flow to the cecum and distal small intestine (correction of intesting) in both groups, whereas it was significantly increased (477%) in the remaining parts of the liver in the liver resection group. CONCLUSIONS: These findings show that reticuloendothelial system activity is deranged in both these groups, which may explain the increased occurrence of bacterial complications observed in corresponding clinical conditions.