Gotardo Érica Martins Ferreira, Ribeiro Gilberto de Almeida, Clemente Thayane Rodrigues Leite, Moscato Camila Henrique, Tomé Renata Bortolin Guerra, Rocha Thalita, Pedrazzoli José, Ribeiro Marcelo Lima, Gambero Alessandra
Érica Martins Ferreira Gotardo, Gilberto de Almeida Ribeiro, Thayane Rodrigues Leite Clemente, Camila Henrique Moscato, Renata Bortolin Guerra Tomé, Thalita Rocha, José Pedrazzoli Jr, Marcelo Lima Ribeiro, Alessandra Gambero, Clinical Pharmacology and Gastroenterology Unit, São Francisco University Medical School, Bragança Paulista, SP 12916-900, Brazil.
World J Gastroenterol. 2014 Apr 21;20(15):4345-52. doi: 10.3748/wjg.v20.i15.4345.
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.
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.
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.
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.
研究在三硝基苯磺酸（TNBS）诱导的大鼠结肠炎模型中，大鼠结肠中铁调素的表达、白细胞介素-6（IL-6）的产生及铁水平。
在大鼠中，我们使用宏观和微观评分系统以及髓过氧化物酶活性测量方法评估了重复给予TNBS诱导的结肠炎的严重程度。通过酶联免疫吸附测定法测量结肠中铁调素、肿瘤坏死因子α（TNF-α）、IL-10和IL-6的水平，分别通过免疫组织化学和普鲁士蓝反应分析铁调素-25的表达和铁沉积情况。通过蛋白质印迹分析评估Stat-3磷酸化。还测量了血液学参数、铁和转铁蛋白水平以及转铁蛋白饱和度。此外，评估了铁、病原体衍生分子和IL-6在HT-29细胞中诱导铁调素表达的能力。
对大鼠重复给予TNBS导致在宏观和微观上均可检测到结肠病变，并使结肠髓过氧化物酶活性升高。结肠炎大鼠结肠表面上皮中的铁调素-25蛋白水平升高（10.2±4.0 pg/mg蛋白对71.0±8.4 pg/mg蛋白，P<0.01）。还观察到IL-6水平升高（8.2±1.7 pg/mg蛋白对14.7±0.7 pg/mg蛋白，P<0.05）、TNF-α水平升高（1.8±1.2 pg/mg蛋白对7.4±2.1 pg/mg蛋白，P<0.05）以及Stat-3磷酸化。在结肠炎大鼠中未检测到铁稳态、铁调素水平和贫血的全身改变。仅在结肠炎期间观察到结肠中的铁沉积。IL-6和脂多糖[一种Toll样受体4（TLR-4）配体]处理后，HT-29细胞中的铁调素基因表达增加。去铁胺、柠檬酸铁和肽聚糖（一种TLR-2配体）无法改变HT-29细胞中铁调素的体外表达。
结肠炎增加了局部铁调素-25的表达，这与IL-6/Stat-3信号通路有关。还观察到局部铁螯合增加，但需要进一步研究以确定这种螯合是对肠道炎症的防御性还是病理性反应。
