Role of metallothionein in murine experimental colitis.

Metallothioneins (MTs) are a family of cysteine-rich low molecular-weight proteins that can act as reactive oxygen species scavengers. Although it is known that the induction of MT expression suppresses various inflammatory disorders, the role of MTs in intestinal inflammation remains unclear. In this study, we investigated the effects of dextran sulfate sodium (DSS) administration in mice with targeted deletions of the MT-I/II genes. Acute colitis was induced by 2% DSS in male MT-I/II double knockout (MT-null) and C57BL/6 (wild-type) mice. The disease activity index (DAI) was determined on a daily basis for each animal, and consisted of a calculated score based on changes in body weight, stool consistency and intestinal bleeding. Histology, colon length, myeloperoxidase (MPO) activity and colonic mRNA expression and the concentration of inflammatory cytokines were evaluated by real-time-PCR and enzyme-linked immunosorbent assay (ELISA). The localization of MTs and macrophages was determined by immunohistological and immunofluorescence staining. To investigate the role of MTs in macrophages, peritoneal macrophages were isolated and their responses to lipopolysaccharide were measured. Following DSS administration, the DAI score increased in a time-dependent manner and was significantly enhanced in the MT-I/II knockout mice. Colonic MPO activity levels and inflammatory cytokines [tumor necrosis factor (TNF)-α, interferon (IFN)-γ and interleukin (IL)-17] production increased following DSS administration, and these increases were significantly enhanced in the MT-I/II knockout mice compared with the wild-type mice. MT-positive cells were detected in the lamina propria and submucosal layer by immunohistochemical and immunofluorescence staining, and were mainly co-localized in F4/80-positive macrophages. The production of inflammatory cytokines (TNF-α, IFN-γ and IL-17) from isolated peritoneal macrophages increased following lipopolysaccharide stimulation, and these increases were significantly enhanced in the macrophages obtained from the MT-I/II knockout mice. These data indicate that MTs play an important role in the prevention of colonic mucosal inflammation in a mouse model of DSS-induced colitis, thus suggesting that endogenous MTs play a protective role against intestinal inflammation.

[1]  M. Stoltenberg,et al.  Metallic gold slows disease progression, reduces cell death and induces astrogliosis while simultaneously increasing stem cell responses in an EAE rat model of multiple sclerosis , 2012, Histochemistry and Cell Biology.

[2]  Kechen Ban,et al.  Protective Role of p70S6K in Intestinal Ischemia/Reperfusion Injury in Mice , 2012, PloS one.

[3]  S. Hauptmann,et al.  Influence of heat shock protein 70 and metallothionein induction by zinc-bis-(DL-hydrogenaspartate) on the release of inflammatory mediators in a porcine model of recurrent endotoxemia. , 1996, Biochemical pharmacology.

[4]  R. Xavier,et al.  Unravelling the pathogenesis of inflammatory bowel disease , 2007, Nature.

[5]  J. Roman,et al.  Zinc supplementation restores PU.1 and Nrf2 nuclear binding in alveolar macrophages and improves redox balance and bacterial clearance in the lungs of alcohol-fed rats. , 2011, Alcoholism, clinical and experimental research.

[6]  P. Coyle,et al.  The Role of Zinc and Metallothionein in the Dextran Sulfate Sodium-Induced Colitis Mouse Model , 2007, Digestive Diseases and Sciences.

[7]  S. Himeno,et al.  Metallothionein is a crucial protective factor against Helicobacter pylori-induced gastric erosive lesions in a mouse model. , 2008, American journal of physiology. Gastrointestinal and liver physiology.

[8]  C. McClain,et al.  Metallothionein overexpression does not protect against inflammatory bowel disease in a murine colitis model. , 2005, Medical science monitor : international medical journal of experimental and clinical research.

[9]  A. Michalska,et al.  Targeting and germ-line transmission of a null mutation at the metallothionein I and II loci in mouse. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[10]  A. Sanz-Medel,et al.  The Stoichiometric Transition from Zn6Cu1-Metallothionein to Zn7-Metallothionein Underlies the Up-regulation of Metallothionein (MT) Expression , 2012, The Journal of Biological Chemistry.

[11]  T. Nishimura,et al.  The Severity of Dextran Sodium Sulfate-Induced Colitis Can Differ Between Dextran Sodium Sulfate Preparations of the Same Molecular Weight Range , 2012, Digestive Diseases and Sciences.

[12]  Y. Naito,et al.  Rosuvastatin, a new HMG-CoA reductase inhibitor, reduces the colonic inflammatory response in dextran sulfate sodium-induced colitis in mice. , 2006, International journal of molecular medicine.

[13]  T. Minami,et al.  Separation of metallothionein isoforms of mouse liver cytosol by capillary zone electrophoresis. , 1998, Talanta.

[14]  K. Tominaga,et al.  Anti-inflammatory effects of IL-17A on Helicobacter pylori-induced gastritis. , 2009, Biochemical and biophysical research communications.

[15]  Carla G. Taylor,et al.  Intestinal Inflammation in Rats Induces Metallothionein in Colonic Submucosa , 2009, Journal of clinical biochemistry and nutrition.

[16]  P. Hunziker,et al.  Amino acid analysis of metallothionein. , 1991, Methods in enzymology.

[17]  B. Vallee,et al.  Flame photometry and spectrometry; principles and applications. , 2006, Methods of biochemical analysis.

[18]  K. Schmid,et al.  Increased expression of metallothionein in inflammatory bowel disease , 2001, Inflammation Research.

[19]  T. Hibi,et al.  TL1A produced by lamina propria macrophages induces Th1 and Th17 immune responses in cooperation with IL‐23 in patients with Crohn's disease , 2010, Inflammatory bowel diseases.

[20]  G. Fantuzzi,et al.  IL-1β-converting enzyme (caspase-1) in intestinal inflammation , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[21]  S. Hanauer,et al.  Inflammatory bowel disease: Epidemiology, pathogenesis, and therapeutic opportunities , 2006, Inflammatory bowel diseases.

[22]  Y. Naito,et al.  Carbon Monoxide Liberated from Carbon Monoxide-Releasing Molecule Exerts an Anti-inflammatory Effect on Dextran Sulfate Sodium-Induced Colitis in Mice , 2011, Digestive Diseases and Sciences.

[23]  H. Verspaget,et al.  Differential mucosal expression of three superoxide dismutase isoforms in inflammatory bowel disease , 2003, The Journal of pathology.

[24]  H. Cooper,et al.  Clinicopathologic study of dextran sulfate sodium experimental murine colitis. , 1993, Laboratory investigation; a journal of technical methods and pathology.

[25]  Philip Smith,et al.  Infection with a Helminth Parasite Prevents Experimental Colitis via a Macrophage-Mediated Mechanism1 , 2007, The Journal of Immunology.

[26]  P. Coyle,et al.  Metallothionein: the multipurpose protein , 2002, Cellular and Molecular Life Sciences CMLS.

[27]  C. Yang,et al.  Immunomodulatory and anti-inflammatory properties of artesunate in experimental colitis. , 2012, Current medicinal chemistry.

[28]  H. Verspaget,et al.  Decrease in two intestinal copper/zinc containing proteins with antioxidant function in inflammatory bowel disease. , 1991, Gut.

[29]  Y. Naito,et al.  Molecular fingerprints of neutrophil-dependent oxidative stress in inflammatory bowel disease , 2007, Journal of Gastroenterology.

[30]  P. Mannon,et al.  Complement-Dependent Injury and Protection in a Murine Model of Acute Dextran Sulfate Sodium-Induced Colitis , 2012, The Journal of Immunology.

[31]  G. Mullin,et al.  Increased oxidative stress and decreased antioxidant defenses in mucosa of inflammatory bowel disease , 1996, Digestive Diseases and Sciences.

[32]  M. Kirkcaldie,et al.  Increased circulating leukocyte numbers and altered macrophage phenotype correlate with the altered immune response to brain injury in metallothionein (MT) -I/II null mutant mice , 2011, Journal of Neuroinflammation.

[33]  I. Moret,et al.  Role of oxidative stress and antioxidant enzymes in Crohn's disease. , 2011, Biochemical Society transactions.

[34]  T. Kimura,et al.  Sensitivity of metallothionein-null mice to LPS/D-galactosamine-induced lethality. , 2001, Biochemical and biophysical research communications.

[35]  C. Xian,et al.  Predisposition to Colonic Dysplasia is Unaffected by Continuous Administration of Insulin-like Growth Factor-1 for Twenty Weeks in a Rat Model of Chronic Inflammatory Bowel Disease , 2000, Growth factors.

[36]  T. Yoshikawa,et al.  Role of metallothionein in coagulatory disturbance and systemic inflammation induced by lipopolysaccharide in mice , 2006, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[37]  P. Mannon,et al.  The fundamental basis of inflammatory bowel disease. , 2007, The Journal of clinical investigation.

[38]  D. Granger,et al.  Impact of Dextran Sulfate Sodium Load on the Severity of Inflammation in Experimental Colitis , 2004, Digestive Diseases and Sciences.

[39]  M. Sato,et al.  Protective role of metallothionein in acute lung injury induced by bacterial endotoxin , 2004, Thorax.

[40]  D. Korenaga,et al.  Impaired antioxidant defense system of colonic tissue and cancer development in dextran sulfate sodium-induced colitis in mice. , 2002, The Journal of surgical research.

[41]  Jay Siddharth,et al.  Host Genetics and Environmental Factors Regulate Ecological Succession of the Mouse Colon Tissue-Associated Microbiota , 2012, PloS one.

[42]  C. Tohyama,et al.  Cytoprotection by Metallothionein Against Gastroduodenal Mucosal Injury Caused by Ethanol in Mice , 2000, Laboratory Investigation.

[43]  S. Shioda,et al.  Effect of interleukin-6 neutralization on CYP3A11 and metallothionein-1/2 expressions in arthritic mouse liver. , 2007, European journal of pharmacology.

[44]  M. Grisham,et al.  Xanthine oxidase and neutrophil infiltration in intestinal ischemia. , 1986, The American journal of physiology.

[45]  Agneta Karlsson,et al.  Acute colitis induced by dextran sulfate sodium progresses to chronicity in C57BL/6 but not in BALB/c mice: correlation between symptoms and inflammation. , 2005, American journal of physiology. Gastrointestinal and liver physiology.