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重組人乙酰膽堿受體α亞基1-210誘導(dǎo)實(shí)驗(yàn)性重癥肌無力模型

發(fā)布時(shí)間:2018-05-04 02:17

  本文選題:人乙酰膽堿受體a亞基1-210 + 實(shí)驗(yàn)性重癥肌無力 ; 參考:《安徽醫(yī)科大學(xué)》2006年碩士論文


【摘要】:背景:重癥肌無力(MG)是一種主要累及神經(jīng)肌肉接頭突觸后膜乙酰膽堿受體(AChR),由乙酰膽堿受體抗體(AchR-ab)介導(dǎo)的自身免疫性疾病。發(fā)病率高達(dá)8~20/10萬,并有逐年上升的趨勢(shì);且致殘率高,嚴(yán)重危害人類健康,尋找有效治療手段一直是研究者們努力的方向。實(shí)驗(yàn)性重癥肌無力(EAMG)是其基礎(chǔ)研究的動(dòng)物模型;構(gòu)建該模型的經(jīng)典方法是由電鰻電器管中提取的純化AchR作為免疫原,電鰻難以捕獲,且提取過程復(fù)雜、費(fèi)用昂貴,因此尋找構(gòu)建該模型的新方法成為當(dāng)務(wù)之急。AChR的主要生物學(xué)功能單位是α亞基,它具有高度的免疫原性,有研究證實(shí)刺激α亞基或α亞基的某些肽段,可以激活A(yù)ChR特異反應(yīng)性T細(xì)胞發(fā)生免疫反應(yīng)。用含有免疫表位的α亞基的某些肽段作為免疫原誘導(dǎo)EAMG模型是近幾十年來研究者們追求的目標(biāo);但可能是由于用作免疫原的肽段多在20個(gè)殘基左右、免疫位點(diǎn)少、免疫原性較弱、須大劑量重復(fù)注射且肌無力存在時(shí)間短暫,不能成為理想的動(dòng)物模型。AchRα亞基的N端細(xì)胞外區(qū)域1-210位殘基,包含了ACh配體的結(jié)合位點(diǎn)和主要免疫原區(qū),,是MG致病的關(guān)鍵區(qū)域,可以推測(cè)該肽段有可能成為有效免疫原。九十年代初Talib就應(yīng)用基因技術(shù)分段克隆了hAChR α1-210片段,并在大腸桿菌中誘導(dǎo)表達(dá);構(gòu)建EAMG模型的知名專家Lennon則以該表達(dá)產(chǎn)物免疫Lewis鼠,結(jié)果成功誘導(dǎo)EAMG模型。隨著分子生物學(xué)的發(fā)展,應(yīng)用分子克隆和表達(dá)技術(shù)重組目的基因的方法已非常成熟;但國(guó)內(nèi)外均未見使用類似方法誘導(dǎo)EAMG模型的報(bào)道。 目的:我們首次參照Talib文獻(xiàn)克隆并在大腸桿菌中誘導(dǎo)表達(dá)了hAChR α 1-210,然后用該表達(dá)產(chǎn)物免疫Lewis鼠誘導(dǎo)EAMG模型,希望為MG的基礎(chǔ)研究提供一個(gè)簡(jiǎn)便、易行的實(shí)驗(yàn)動(dòng)物模型。
[Abstract]:Background: myasthenia gravis (MG) is an autoimmune disease mediated by acetylcholine receptor (ache) receptor (ACHR), which mainly involves the postsynaptic membrane of neuromuscular junctions. The incidence rate is as high as 820 / 100, 000, and has a rising trend year by year, and the rate of disability is high, which seriously endangers human health. Finding effective treatment methods has been the direction of researchers' efforts. Experimental myasthenia gravis (EAMG) is an animal model for its basic research. The classical method to construct this model is to use purified AchR extracted from electric eel tube as immunogen, which is difficult to capture, and the extraction process is complex and expensive. Therefore, searching for a new method to construct this model has become an urgent task. The main biological functional unit of AChR is 偽 subunit, which has high immunogenicity. Some studies have confirmed that stimulating 偽 subunit or certain peptide segments of 偽 subunit. AChR specific reactive T cells can be activated to develop immune response. Using some peptides of 偽 subunit containing immune epitopes as immunogen to induce EAMG model has been pursued by researchers in recent decades, but this may be due to the fact that most of the peptides used as immune epitopes are about 20 residues and there are few immune sites. The immunogenicity is weak, the large dose of repeated injection and the short duration of myasthenia can not be an ideal animal model. The extracellular region of the N-terminal region of the AchR 偽 subunit is 1-210 residues, which contains the binding site and the main immunogen region of the ACh ligand. It can be inferred that the peptide may become an effective immunogen. In the early 1990s, Talib cloned the fragment of hAChR 偽 1-210 and induced expression in Escherichia coli, and Lennon, a well-known expert who constructed the EAMG model, immunized Lewis mice with the expression product, and the EAMG model was successfully induced. With the development of molecular biology, the methods of molecular cloning and expression to recombine the target gene have been very mature, but no similar methods have been reported at home and abroad to induce EAMG model. Objective: we cloned and induced the expression of hAChR 偽 1-210 in Escherichia coli with reference to Talib for the first time, and then immunized Lewis mice with this expression product to induce EAMG model, hoping to provide a simple and easy experimental animal model for the basic study of MG.
【學(xué)位授予單位】:安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】:碩士
【學(xué)位授予年份】:2006
【分類號(hào)】:R746.1;R-332

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