【摘要】：巨噬細(xì)胞是先天免疫系統(tǒng)的重要組成部分,在免疫調(diào)節(jié)中充當(dāng)重要角色。很多研究從巨噬細(xì)胞活化及調(diào)節(jié)細(xì)胞因子分泌、遷移等角度,研究腸道炎癥的發(fā)病機(jī)理。表皮生長(zhǎng)因子(EGF, epidermal growth factor)信號(hào)通路激活后可促進(jìn)細(xì)胞生長(zhǎng),但是巨噬細(xì)胞中EGF信號(hào)通路的作用尚未闡明。本課題組一直從事腸道炎癥中EGF信號(hào)通路作用的基礎(chǔ)研究。本課題即在前期研究的基礎(chǔ)上,探索巨噬細(xì)胞中EGF受體(EGFR, epidermal growth factor receptor)激活對(duì)細(xì)胞因子及腸道炎癥的調(diào)節(jié)作用。 本課題研究分為以下三個(gè)部分： 第一部分,首先在體外實(shí)驗(yàn)中證明了在巨噬細(xì)胞中抑制EGFR激酶活性能夠上調(diào)脂多糖(LPS, lipopolysaccharide)和干擾素-γ(IFN-γ,interferon-γ)刺激下的炎性細(xì)胞因子,如腫瘤壞死因子(TNF, tumor necrosis factor)和白介素-10(IL-10)的表達(dá),其與p38及核因子-κB (NF-κB)表達(dá)增加相關(guān)。其次在巨噬細(xì)胞EGFR表達(dá)缺陷小鼠模型中,發(fā)現(xiàn)巨噬細(xì)胞中敲除EGFR能夠上調(diào)LPS刺激的炎性細(xì)胞因子表達(dá)。 第二部分,利用葡聚糖硫酸鈉(DSS, dextran sulfate sodium)誘發(fā)的小鼠結(jié)腸炎模型,發(fā)現(xiàn)巨噬細(xì)胞中敲除EGFR能夠促進(jìn)結(jié)腸炎緩解并增強(qiáng)組織修復(fù)。在炎癥急性期和修復(fù)期中,巨噬細(xì)胞中敲除EGFR后對(duì)TNF-α和IL-10具有不同的調(diào)節(jié)作用。隨后驗(yàn)證了在巨噬細(xì)胞EGFR表達(dá)缺陷小鼠模型中,IL-10參與介導(dǎo)了EGFR對(duì)巨噬細(xì)胞免疫應(yīng)答的調(diào)節(jié)作用,巨噬細(xì)胞EGFR表達(dá)缺陷小鼠與對(duì)照組相比,IL-10表達(dá)水平在結(jié)腸炎急性期及恢復(fù)期均持續(xù)增加,而TNF表達(dá)水平在急性期增加,在恢復(fù)期減少,Anti-IL-10中和抗體能夠消除上述作用�？梢�(jiàn)IL-10能夠抑制TNF生成,從而對(duì)巨噬細(xì)胞EGFR表達(dá)缺陷小鼠DSS結(jié)腸炎起到保護(hù)作用。 第三部分,在巨噬細(xì)胞EGFR表達(dá)缺陷小鼠DSS結(jié)腸炎中,發(fā)現(xiàn)結(jié)腸上皮細(xì)胞增殖增加,組織中ki67陽(yáng)性細(xì)胞增加,同時(shí)結(jié)腸巨噬細(xì)胞中及結(jié)腸組織中環(huán)氧合酶-2(cyclooxygenase-2, COX-2) mRNA水平增加。利用免疫熒光法檢測(cè)潰瘍性結(jié)腸炎(ulcerative colitis, UC)患者結(jié)腸鏡活檢標(biāo)本,發(fā)現(xiàn)EGFR磷酸化水平及人巨噬細(xì)胞標(biāo)志物CD68表達(dá)水平增加。 綜上所述,可以得出如下結(jié)論：在巨噬細(xì)胞中敲除EGFR后能夠增加致炎細(xì)胞因子TNF及抗炎細(xì)胞因子IL-10的生成,IL-10生成增加會(huì)抑制TNF的生成,從而對(duì)腸道炎癥起到保護(hù)作用。
[Abstract]:Macrophages are an important part of innate immune system and play an important role in immune regulation. Many studies have studied the pathogenesis of intestinal inflammation from the point of view of macrophage activation and regulation of cytokine secretion and migration. The activation of epidermal growth factor (EGF, epidermal growth factor) signaling pathway can promote cell growth, but the role of EGF signaling pathway in macrophages has not been clarified. Our research group has been engaged in the basic research of EGF signaling pathway in intestinal inflammation. On the basis of previous research, this study explored the regulatory effect of EGF receptor (EGFR, epidermal growth factor receptor) activation on cytokines and intestinal inflammation in macrophages. This study is divided into the following three parts: in the first part, it was proved in vitro that inhibition of EGFR kinase activity in macrophages can up-regulate lipopolysaccharide (LPS, lipopolysaccharide) and interferon-gamma (IFN- gamma). The expression of inflammatory cytokines stimulated by interferon- 緯, such as tumor necrosis factor (TNF, tumor necrosis factor) and IL-10 (IL-10), was correlated with the increased expression of p38 and nuclear factor-kappa B (NF- kappa B). Secondly, in macrophage EGFR expression deficient mouse model, it was found that knockout EGFR in macrophages could up-regulate the expression of LPS-stimulated inflammatory cytokines. In the second part, using the mouse colitis model induced by dextran sodium sulfate (DSS, dextran sulfate sodium), it was found that knockout of EGFR in macrophages could promote the remission of colitis and enhance tissue repair. In the acute phase and repair stage of inflammation, the knockout of EGFR in macrophages has different regulatory effects on TNF- 偽 and IL-10. Then it was verified that IL-10 was involved in the regulation of macrophage immune response by EGFR in macrophage EGFR expression deficient mouse model. Compared with the control group, macrophage EGFR expression deficient mice were involved in the regulation of macrophage immune response. The expression of IL-10 continued to increase in both acute and convalescent colitis, while the expression of TNF increased in acute phase and decreased in convalescent phase. Anti-IL-10 neutralizing antibody could eliminate the above effects. It can be seen that IL-10 can inhibit TNF production, thus protecting DSS colitis in mice with macrophage EGFR expression deficiency. In the third part, in DSS colitis of mice with macrophage EGFR expression deficiency, it was found that the proliferation of colon epithelial cells increased, ki67 positive cells increased, and cycloxygenase-2 (cyclooxygenase-2,) was found in colon macrophages and colon tissues. COX-2) mRNA level increased. Colonoscopy biopsies from patients with ulcerative colitis (ulcerative colitis, UC) were detected by immunofluorescence. It was found that the phosphorylation of EGFR and the expression of CD68, a human macrophage marker, were increased. In conclusion, the following conclusions can be drawn: knockout of EGFR in macrophages can increase the production of inflammatory cytokine TNF and anti-inflammatory cytokine IL-10, and the increase of IL-10 production will inhibit the production of TNF. Thus, it plays a protective role in intestinal inflammation.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R574.62

【參考文獻(xiàn)】

相關(guān)期刊論文 前4條

1 ;Cellular and Molecular Immunopathogenesis of Ulcerative Colitis[J];Cellular & Molecular Immunology;2006年01期

2 Jesus K Yamamoto-Furusho;;Innovative therapeutics for inflammatory bowel disease[J];World Journal of Gastroenterology;2007年13期

3 Emilio Cuadrado;Marta Alonso;Maria Dolores de Juan;Pilar Echaniz;Juan Ignacio Arenas;;Regulatory T cells in patients with inflammatory bowel diseases treated with adacolumn granulocytapheresis[J];World Journal of Gastroenterology;2008年10期

4 Avi Levin;Oren Shibolet;;Toll-like receptors in inflammatory bowel disease-stepping into uncharted territory[J];World Journal of Gastroenterology;2008年33期

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本文編號(hào)：2496850