Iron is an essential metallic microelement for life. However, iron overload is toxic. The liver serves an important role as a storehouse for iron in the body. About 20–25 mg of iron is required each day for hemoglobin synthesis. To maintain iron homeostasis, transferrin and transferrin receptors are primarily involved in the uptake of iron into hepatocytes, ferritin in its storage, and ferroportin in its export. Moreover, hepcidin controls ferroportin and plays a central role in the iron metabolism. Excess “free” reactive iron produces damaging free radicals via Fenton or Harber-Weiss reactions. Produced free radicals attack cellular proteins, lipids and nucleic acid. Several detoxification system and anti-oxidant defense mechanisms exist to prevent cellular damage by free radicals. Excessive free radicals can lead to hepatocellular damage, liver fibrosis, and hepatocarcinogenesis.
Non-steroidal anti-inflammatory drugs are the most commonly prescribed drugs for arthritis, inflammation, and cardiovascular protection. However, they cause gastrointestinal complications. The pathophysiology of these complications has mostly been ascribed to non-steroidal anti-inflammatory drugs’ action on the cyclooxygenase inhibition and the subsequent prostaglandin deficiency. However, recent clinical demonstrated the prevalence of non-steroidal anti-inflammatory drugs-induced small intestinal mucosal injury is more often than previously expected. In this review, we discuss the defense mechanisms of stomach, and the pathophysiology of non-steroidal anti-inflammatory drugs-induced injury of stomach and small intestine, especially focused on non-steroidal anti-inflammatory drugs’ action on mitochondria.
The pathophysiology of inflammatory bowel disease involves excessive immune effects of inflammatory cells against gut microbes. In genetically predisposed individuals, these effects are considered to contribute to the initiation and perpetuation of mucosal injury. Oxidative stress is a fundamental tissue-destructive mechanisms that can occur due to the reactive oxygen species and reactive nitrogen metabolites which are released in abundance from numerous inflammatory cells that have extravasated from lymphatics and blood vessels to the lamina propria. This extravasation is mediated by interactions between adhesion molecules including mucosal addressin cell adhesion molecule-1 and vascular cell adhesion molecule-1 on the surface of lymphocytes or neutrophils and their ligands on endothelial cells. Thus, reactive oxygen species and adhesion molecules play an important role in the development of inflammatory bowel disease. The present review focuses on the involvement of oxidative stress and adhesion molecules, in particular mucosal addressin cell adhesion molecule-1, in inflammatory bowel disease.
Recent studies using small bowel endoscopy revealed that non-steroidal anti-inflammatory drugs including low-dose aspirin, can often induce small bowel injury. Non-steroidal anti-inflammatory drugs-induced small bowel mucosal injury involves various factors such as enterobacteria, cytokines, and bile. Experimental studies demonstrate that both mitochondrial disorders and inhibition of cyclooxygenases are required for development of non-steroidal anti-inflammatory drugs-induced small bowel injury. Mitochondrion is an organelle playing a central role in energy production in organisms. Many non-steroidal anti-inflammatory drugs directly cause mitochondrial disorders, which are attributable to uncoupling of oxidative phosphorylation induced by opening of the mega channel called mitochondrial permeability transition pore on the mitochondrial membrane by non-steroidal anti-inflammatory drugs. Bile acids and tumor necrosis factor-α also can open the permeability transition pore. The permeability transition pore opening induces the release of cytochrome c from mitochondrial matrix into the cytosol, which triggers a cascade of events that will lead to cell death. Therefore these mitochondrial disorders may cause disturbance of the mucosal barrier function and elevation of the small bowel permeability, and play particularly important roles in early processes of non-steroidal anti-inflammatory drugs-induced small bowel injury. Although no valid means of preventing or treating non-steroidal anti-inflammatory drugs-induced small bowel injury has been established, advances in mitochondrial studies may bring about innovation in the prevention and treatment of this kind of injury.
Oxidative stress is a major cause of the gastrointestinal damage under physical or psychological stress. Ghrelin exhibits gastroprotective effects and they are supposed to be derived from antioxidant effects. In gastroduodenal mucosal injury, the plasma ghrelin levels increase in response to the demand for gastroduodenal cytoprotection. However, in the condition of Helicobacter pylori-induced gastric mucosal severe atrophy, the plasma ghrelin concentration shifted to lower levels. In diabetic gastroparesis, the regulation of ghrelin secretion is impaired with vagal nerve dysfunction. Selective ghrelin agonist is expected to represent a new class of prokinetic agent. In addition, the plasma ghrelin levels are also enhanced by systemic oxidative stress, and ghrelin exhibits antioxidant effects in many organs, such as heart, pancreas, and lung. This suggests that ghrelin would be an important player as a sensor of systemic oxidative stress.
Heme oxygenase-1 (HO-1) is the rate-limiting enzyme in the catabolism of heme, followed by production of biliverdin, free iron and carbon monoxide (CO). HO-1 is a stress-responsive protein induced by various oxidative agents. Recent studies demonstrate that the expression of HO-1 in response to different inflammatory mediators may contribute to the resolution of inflammation and has protective effects in several organs against oxidative injury. Although the mechanism underlying the anti-inflammatory actions of HO-1 remains poorly defined, both CO and biliverdin/bilirubin have been implicated in this response. In the gastrointestinal tract, HO-1 is shown to be transcriptionally induced in response to oxidative stress, preconditioning and acute inflammation. Recent studies suggest that the induction of HO-1 expression plays a critical protective role in intestinal damage models induced by ischemia-reperfusion, indomethacin, lipopolysaccharide-associated sepsis, trinitrobenzene sulfonic acid, and dextran sulfate sodium, indicating that activation of HO-1 may act as an endogenous defensive mechanism to reduce inflammation and tissue injury in the gastrointestinal tract. In addition, CO derived from HO-1 is shown to be involved in the regulation in gastro-intestinal motility. These in vitro and in vivo data suggest that HO-1 may be a novel therapeutic target in patients with gastrointestinal diseases.
Lipopolysaccharide-stimulated leukocytes secrete proinflammatory cytokines including tumor necrosis factor-α and interleukin-12. Over-activation of host defense systems may result in severe tissue damage and requires regulation. Granulocyte colony-stimulating factor and interleukin-10 are candidate cytokines for inducing tolerance to lipopolysaccharide re-stimulation. We compared cytokines secreted by lipopolysaccharide-stimulated blood cells from patients who had survived gram negative bacterial pneumonia (Pseudomonas aeruginosa, Escherichia coli or Proteus mirabilis, n = 26) and age-matched healthy volunteers (n = 18). Interleukin-12p70 and tumor necrosis factor-α expression was significantly lower in patients (p = 0.0039 and p<0.001) compared to healthy controls, while granulocyte colony-stimulating factor production was markedly higher in patients (p<0.001). Levels of interleukin-10 were comparable. Granulocyte colony-stimulating factor expression was inversely correlated with interleukin-12p70 (R = −0.71, p<0.001) and tumor necrosis factor-α (R = −0.64, p<0.001) expression; interleukin-10 showed no significant correlation. In unstimulated leukocytes from patients, cAMP levels were significantly raised (p = 0.020) and were correlated inversely with interleukin-12p70 levels (R = −0.81, p<0.001) and directly with granulocyte colony-stimulating factor (R = 0.72, p = 0.0020), matrix metalloproteinase-9 (R = 0.67, p = 0.0067) and interleukin-10 (R = 0.54, p = 0.039) levels. Our results demonstrate that granulocyte colony-stimulating factor production by lipopolysaccharide-stimulated leukocytes is a useful indicator of tolerance induction in surviving pneumonia patients and that measuring cAMP in freshly isolated leukocytes may also be clinically significant.
Liver steatosis is associated with organ dysfunction after hepatic resection and transplantation which may be caused by hepatic ischemia/reperfusion injury. The aim of the current study was to determine the precise mechanism leading to hepatocyte apoptosis after steatotic liver ischemia/reperfusion. Using a murine model of partial hepatic ischemia for 90 min, we examined the levels and pathway of apoptosis, and the peroxynitrite expression, serum alanine aminotransferase levels, and liver histology 1 and 4 h after reperfusion. In the steatotic liver, the peroxynitrite expression increased after ischemia/reperfusion. Significant hepatocyte apoptosis in the steatotic liver was seen after reperfusion, caused by upregulation of cleaved caspases 9 and 3, but not caspase 8. Serum alanine aminotransferase levels were elevated and histological examination revealed severe liver injury in the steatotic liver 4 h after reperfusion. In mice treated with aminoguanidine, ischemia/reperfusion-induced increases in serum alanine aminotransferase levels and apoptosis were significantly reduced in steatotic liver compared with mice treated with phosphate buffered saline. Survival of mice with steatotic livers significantly improved by treatment with aminoguanidine. Our data suggested that the steatotic liver is vulnerable to hepatic ischemia/reperfusion, leading to significant hepatocyte apoptosis by the mitochondrial permeability transition, and thereby resulting in organ dysfunction.
Prostaglandin E2 plays an important role in the maintenance of gastric mucosal integrity. The level of biologically active prostaglandin E2 in the tissue is regulated by the balanced expression of its synthetic enzymes, such as cyclooxygenase, and its catabolic enzyme, 15-hydroxyprostaglandin dehydrogenase. We examined the effect of rebamipide, a mucoprotective drug, on prostaglandin E2 production and metabolism in the gastric tissue and its effect on indomethacin-induced gastric mucosal injury in mice. Rebamipide suppressed indomethacin-induced gastric mucosal injury. Suppressive effect of rebamipide on indomethacin-induced gastric mucosal injury was also observed in cyclooxygenase-2-knockout mice. The mice that were treated with rebamipide showed a 2-fold increase in cyclooxygenase-2 mRNA expression in the gastric tissue, whereas 15-hydroxyprostaglandin dehydrogenase mRNA expression markedly decreased as compared to vehicle-treated control mice. Rebamipide did not affect the expression of cyclooxygenase-1 in the gastric tissue. Rebamipide did not increase prostaglandin E2 production in the gastric tissue; however, it induced a 1.4-fold increase in the concentration of prostaglandin E2 in the gastric tissue as compared to vehicle-treated control mice. These results suggest that the suppressive effect of rebamipide on non-steroidal anti-inflammatory drugs-induced gastric mucosal injury can be attributed to reduced 15-hydroxyprostaglandin dehydrogenase expression, which increases the prostaglandin E2 concentration in the gastric tissue.
TANK-binding kinase 1 (TBK1) regulates the interferon regulatory factor (IRF) 3 and IRF7 activation pathways by double strand RNA (dsRNA) via Toll-like receptor (TLR) 3 and by lipopolysaccharide (LPS) via TLR4. Rebamipide is useful for treating inflammatory bowel disease (IBD). Although IBD is associated with nuclear factor-κB (NF-κB), any association with the TBK1-IRF pathway remains unknown. How rebamipide affects the TBK1-IRF pathway is also unclear. We analyzed the relationship between IBD (particularly ulcerative colitis; UC) and the TLR-TBK1-IRF3/7 pathway using human colon tissue, a murine model of colitis and human colonic epithelial cells. Inflamed colonic mucosa over-expressed TKB1, NAP1, IRF3, and IRF7 mRNA compared with normal mucosa. TBK1 was mainly expressed in inflammatory epithelial cells of UC patients. The expression of TBK1, IRF3, IRF7, IFN-α and IFN-β mRNA was suppressed in mice given oral dextran sulfate-sodium (DSS) and daily rectal rebamipide compared with mice given only DSS. Rebamipide reduced the expression of TBK1, IRF3 and IRF7 mRNA induced by LPS/dsRNA, but not of NF-κB mRNA in colonic epithelial cells. Rebamipide might suppress the TLR-TBK1 pathway, resulting in IRF3/7-induction of IFN-α/β and inflammatory factors. TBK1 is important in the induction of inflammation in patients with UC. If rebamipide represses the TLR-TBK1 pathway, then rectal administration should suppress inflammation of the colonic mucosa in patients with UC.
Bach1 is a transcriptional repressor of heme oxygenase-1 (HO-1, a.k.a. HSP-32), which is an inducible enzyme and has anti-oxidation/anti-inflammatory properties shown in various models of organ injuries. Since oxidative stress plays a pivotal role in the pathogenesis of nonalcoholic steatohepatitis (NASH), HO-1 induction would be expected to prevent the development of NASH. In this study, we investigated the influence of Bach1 ablation in mice on the progression of NASH in methionine-choline deficient (MCD) diet model. Bach1 ablation resulted in significant induction of HO-1 mRNA and its activity in the liver. When fed MCD diet, Bach1−/− mice exhibited negligible hepatic steatosis compared to pronounced steatohepatitis in wild type mice with 6-fold increase in hepatic triglyceride content. Whereas feeding of MCD diet decreased mRNA expressions of peroxisome proliferator-activated receptor (PPAR) α and microsomal triglyceride transfer protein (MTP) in wild type mice, there were no change in Bach1−/− mice. In addition, hepatic concentration of malondialdehyde (MDA), a biomarker for oxidative stress as well as plasma alanine aminotransferase (ALT) was significantly lower in Bach1−/− mice. These findings suggest that Bach1 ablation exerts hepatoprotective effect against steatohepatitis presumably via HO-1 induction and may be a potential therapeutic target.
Although alcohol abuse is the most common cause of liver cirrhosis in the United States, the enhancing effects of alcohol on the long-term prognosis of hepatitis C virus (HCV) related liver cirrhosis has not been clarified. To investigate how alcohol abuse influences the prognosis of hepatitis virus related liver cirrhosis, we studied 716 Japanese patients. Cumulative survival and hepatocellular carcinoma (HCC) development rates were analyzed in alcohol abusive, cirrhotic patients with or without hepatitis virus infection. Patients who abused alcohol were younger (p<0.0001) than HCV infected, non-abusive patients. The overall survival rate among patients with alcoholic cirrhosis (Al group), HCV related cirrhosis (HCV group), and HCV infected + alcoholic cirrhosis (HCV + Al group), showed no significant differences, although the 10-year cumulative survival rate of Al group was the highest of the three groups. The HCC development rate of Al group was the lowest. In addition, alcohol abuse decreased the survival rates of HCV group in the early stage with no HCC (p = 0.0028). In conclusion, alcohol abuse might affect the progression of liver damage in HCV infected patients with liver cirrhosis in the early stage, although the influence of alcohol abuse on the long term prognosis seems to be rather small.
The beneficial effects by peroxisome proliferator-activated receptor-γ (PPAR-γ) on gastric injury induced by ischemia-reperfusion have been confirmed, however, the precise mechanism of its cytoprotection is not elucidated thoroughly. The aim of the present study was to determine the gastric localization of PPAR-γ expression in the rat gastric mucosa, and to clarify the mechanism of its cytoprotective properties. The gastric expression of PPAR-γ was confirmed by RT-PCR and western blot, and localized on gastric epithelial cells. The protective effect of PPAR-γ ligands, pioglitazone or 15-deoxy-Δ12,14-prostaglandin J2, on gastric ischemia-reperfusion injury was reversed by the co-administration with PPAR-γ antagonist. The gastric expression of tumor necrosis factor-α and cytokine-induced neutrophil chemoattractant-1 increased significantly in rats treated ischemia-reperfusion, and these increases were significantly inhibited by treatment with pioglitazone. Among the 1,032 probes, 18 probes were up-regulated at least 1.5-fold, 17 were down-regulated at least 1.5-fold by pioglitazone. The network including calnexin, endoplasmic reticulum stress protein, heat shock proteins, and proteasome genes was induced by pioglitazone treatment. In conclusion, activation of gastric epithelial PPAR-γ receptor by its ligands may represent a novel therapeutic approach for gastric inflammation via up-regulation of heat shock proteins and endoplasmic reticulum-related proteins.
Previous studies have shown that activated neutrophils and their myeloperoxidase (MPO)-derived products play a crucial role in the pathogenesis of non-steroidal anti-inflammatory drug (NSAID)-related small intestinal injury. The aim of the present study is to identify dihalogenated proteins in the small intestine on indomethacin administration. Intestinal damage was induced by subcutaneous administration of indomethacin (10 mg/kg) in male Wistar rats, and the severity of the injury was evaluated by measuring the area of visible ulcerative lesions. Tissue-associated MPO activity was measured in the intestinal mucosa as an index of neutrophil infiltration. The dihalogenated proteins were separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) using novel monoclonal antibodies against dibromotyrosine (DiBrY), and they were identified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) peptide mass fingerprinting and a Mascot database search. Single administration of indomethacin elicited increased ulcerative area and MPO activity in the small intestine. 2D-PAGE showed an increased level of DiBrY-modified proteins in the indomethacin-induced injured intestinal mucosa and 6 modified proteins were found. Enolase-1 and albumin were found to be DiBrY modified. These proteins may be responsible for the development of neutrophil-associated intestinal injury induced by indomethacin.