ABSTRACT
AIM: To investigate hepcidin expression, interleukin-6 (IL-6) production and iron levels in the rat colon in the presence of trinitrobenzene sulfonic acid (TNBS)-induced colitis. METHODS: In rats, we evaluated the severity of colitis induced by repeated TNBS administration using macroscopic and microscopic scoring systems and myeloperoxidase activity measurements. The colonic levels of hepcidin, tumor necrosis factor alpha (TNF-α), IL-10 and IL-6 were measured by Enzyme-Linked Immunosorbent Assay, and hepcidin-25 expression and iron deposition were analyzed by immunohistochemistry and the Prussian blue reaction, respectively. Stat-3 phosphorylation was assessed by Western blot analysis. Hematological parameters, iron and transferrin levels, and transferrin saturation were also measured. Additionally, the ability of iron, pathogen-derived molecules and IL-6 to induce hepcidin expression in HT-29 cells was evaluated. RESULTS: Repeated TNBS administration to rats resulted in macroscopically and microscopically detectable colon lesions and elevated colonic myeloperoxidase activity. Hepcidin-25 protein levels were increased in colonic surface epithelia in colitic rats (10.2 ± 4.0 pg/mg protein vs 71.0 ± 8.4 pg/mg protein, P < 0.01). Elevated IL-6 levels (8.2 ± 1.7 pg/mg protein vs 14.7 ± 0.7 pg/mg protein, P < 0.05), TNF-α levels (1.8 ± 1.2 pg/mg protein vs 7.4 ± 2.1 pg/mg protein, P < 0.05) and Stat-3 phosphorylation were also observed. Systemic alterations in iron homeostasis, hepcidin levels and anemia were not detected in colitic rats. Iron deposition in the colon was only observed during colitis. Hepcidin gene expression was increased in HT-29 cells after IL-6 and lipopolysaccharide [a toll-like receptor 4 (TLR-4) ligand] treatment. Deferoxamine, ferric citrate and peptidoglycan (a TLR-2 ligand) were unable to alter the in vitro expression of hepcidin in HT-29 cells. CONCLUSION: Colitis increased local hepcidin-25 expression, which was associated with the IL-6/Stat-3 signaling pathway. An increase in local iron sequestration was also observed, but additional studies are needed to determine whether this sequestration is a defensive or pathological response to intestinal inflammation.
Subject(s)
Colitis/chemically induced , Colitis/metabolism , Colon/metabolism , Hepcidins/metabolism , Trinitrobenzenesulfonic Acid/chemistry , Animals , Colon/drug effects , Enzyme-Linked Immunosorbent Assay , Gene Expression Profiling , Gene Expression Regulation , Interleukin-10/metabolism , Interleukin-6/metabolism , Iron/metabolism , Male , Peroxidase/metabolism , Phosphorylation , Rats , Rats, Wistar , Tumor Necrosis Factor-alpha/metabolismABSTRACT
OBJECTIVE: Infliximab is a monoclonal anti-TNF-α antibody that is used therapeutically to treat Crohn's disease (CD). High levels of pro-inflammatory cytokines, especially TNF-α, have been observed in the gastrointestinal tract of CD patients and were associated with alterations in the mesenteric adipose tissue, which also contributed to the high levels of adipokine release. The authors used a rat model of colitis that produces mesenteric adipose tissue alterations that are associated with intestinal inflammation to study the effects that infliximab treatment has on adipokine production, morphological alterations in adipose tissue and intestinal inflammation. MATERIAL AND METHODS: The ability of infliximab to neutralize rat TNF-α was evaluated in vitro using U937 cells. Colitis was induced by repeated intracolonic trinitrobenzene sulfonic acid instillations and was evaluated by macroscopic score, histopathological analysis, myeloperoxidase activity, TNF-α and IL-10 expression as well as iNOS (inducible NO synthase) expression and JNK phosphorylation in colon samples. The alterations in adipose tissue were assessed by TNF-α, IL-10, leptin, adiponectin and resistin levels as well as adipocyte size and peroxisome proliferator-activated receptor (PPAR)-γ expression. RESULTS: Infliximab treatment controlled intestinal inflammation, which reduced lesions and neutrophil infiltration. Inflammatory markers, such as iNOS expression and JNK phosphorylation, were also reduced. In mesenteric adipose tissue, infliximab increased the production of IL-10 and resistin, which was associated with the restoration of adipocyte morphology and PPAR-γ expression. CONCLUSIONS: Our results suggest that infliximab could contribute to the control of intestinal inflammation by modifying adipokine production by mesenteric adipose tissue.
