本文選題：GPR97 + 基因敲除��； 參考：《華東師范大學(xué)》2014年碩士論文

【摘要】：G蛋白偶聯(lián)受體(G protein-coupled receptors, GPCRs),又稱為七次跨膜受體,是目前已知的細(xì)胞表面第一大類受體家族,在將細(xì)胞外信號(hào)傳入到細(xì)胞內(nèi)起著重要的作用。GPCR在人體中數(shù)量極多,其基因數(shù)量約占基因組的2%,介導(dǎo)多種生理學(xué)反應(yīng),發(fā)生突變會(huì)導(dǎo)致多種疾病,是現(xiàn)在病理學(xué)和藥物學(xué)集中研究的焦點(diǎn)。GPR97屬于G蛋白偶聯(lián)受體2類家族,包含549個(gè)氨基酸殘基和一段較長(zhǎng)的胞外N段,屬于孤兒粘附性G蛋白偶聯(lián)受體。目前其功能尚不明確。由于已知的粘附性GPCRs都與免疫功能存在較大關(guān)系,因此GPR97也可能具有免疫調(diào)節(jié)功能。多發(fā)性硬化癥(Multiple sclerosis, MS)是一種中樞神經(jīng)系統(tǒng)的自身免疫性疾病,是導(dǎo)致青壯年人群非創(chuàng)傷性殘疾的首要疾病。其發(fā)病原因與多種因素有關(guān),包括遺傳、環(huán)境和自身免疫等,因此,盡管研究較多,但尚未找到有效的治愈手段。EAE (Experimental Autoimmune Encephalomyelitis)是常用的研究MS的動(dòng)物模型,在發(fā)病機(jī)理和臨床病理方面與MS有很多相似之處。通常采用MOG多肽免疫小鼠的方法制備EAE模型,已經(jīng)被廣泛應(yīng)用于MS的發(fā)病機(jī)理及尋找有效治療方法的研究之中。樹(shù)突狀細(xì)胞(Dendritic cell)是人體內(nèi)最重要的抗原遞呈細(xì)胞,是連接固有免疫和獲得性免疫的重要橋梁。未成熟的DC主要行使抗原吞噬和加工功能,成熟的DC高表達(dá)MHC分子和共刺激分子如CD40、CD86等,并能通過(guò)分泌不同的細(xì)胞因子誘導(dǎo)初始Th細(xì)胞向不同的細(xì)胞方向進(jìn)行分化。大量的研究表明,DC細(xì)胞可能與EAE的發(fā)病有密切的關(guān)系。GPR97基因敲除小鼠是研究GPR97基因功能的有力工具。我們檢測(cè)了GPR97基因敲除小鼠脾細(xì)胞中的免疫細(xì)胞及其亞群。結(jié)果顯示,GPR97基因敲除小鼠脾細(xì)胞中的T細(xì)胞數(shù)量略低于野生型小鼠,而NK細(xì)胞、CD4+T細(xì)胞及Thl和Th17型細(xì)胞數(shù)量均略高于野生型小鼠。樹(shù)突狀細(xì)胞的檢測(cè)結(jié)果表明,GPR97基因敲除小鼠脾臟中的樹(shù)突狀細(xì)胞數(shù)量顯著高于野生型小鼠,而且具有更高的表面分子的表達(dá)。為了研究GPR97在EAE發(fā)病進(jìn)程中的作用,我們利用GPR97基因敲除的小鼠為研究對(duì)象,采用MOG多肽免疫法建立了EAE模型,通過(guò)分析發(fā)病小鼠的臨床評(píng)分、脊髓組織的病理切片及相關(guān)免疫學(xué)指標(biāo)的檢測(cè)來(lái)判定GPR97基因敲除對(duì)EAE小鼠發(fā)病的影響作用。結(jié)果顯示,與野生型小鼠的EAE模型相比,GPR97基因敲除小鼠的EAE模型臨床評(píng)分更高,中樞神經(jīng)系統(tǒng)中炎癥細(xì)胞浸潤(rùn)現(xiàn)象更加嚴(yán)重,脾臟中Th1和Th17細(xì)胞數(shù)量更多,炎癥因子TNF-α、IL-17A和IL-6表達(dá)水平更高,MOG特異性T細(xì)胞數(shù)量更多,初步證明了GPR97基因敲除的小鼠EAE的發(fā)病程度更加嚴(yán)重。為了進(jìn)一步探討GPR97影響EAE發(fā)病可能的機(jī)制,我們以DC細(xì)胞為靶細(xì)胞,檢測(cè)了GPR97基因敲除對(duì)DC細(xì)胞的細(xì)胞因子分泌譜、表面分子的表達(dá)、誘導(dǎo)Th的分化、抗原遞呈等生物學(xué)功能的影響。RT-PCR和ELISA檢測(cè)結(jié)果顯示,GPR97基因敲除的DC能分泌更多IL-6、TGF-β和IL-12,而IL-10的分泌量減少；GPR97基因敲除的DC細(xì)胞主要誘導(dǎo)Th細(xì)胞向Th1和Th17細(xì)胞分化；GPR97基因敲除的DC細(xì)胞表面分子CD40和CD86的表達(dá)明顯增多；GPR97基因敲除的DC細(xì)胞具有更強(qiáng)的刺激MOG特異性T細(xì)胞增殖的能力。這些結(jié)果表明GPR97基因敲除增強(qiáng)了DC細(xì)胞的生物學(xué)功能。因此,本研究結(jié)果表明GPR97基因與EAE小鼠的發(fā)病有關(guān),其可能的機(jī)制是GPR97基因敲除增強(qiáng)了DC的免疫功能。該研究為探討以GPR97為靶點(diǎn)進(jìn)行MS的治療奠定了一定的理論和實(shí)驗(yàn)基礎(chǔ)。
[Abstract]:The G protein coupling receptor (G protein-coupled receptors, GPCRs), also known as the seven transmembrane receptor, is the first known family of receptors on the surface of the cell. It plays an important role in the introduction of extracellular signals into cells, and the number of.GPCR is very large in the human body. The number of genes is about 2% of the genome, which mediates a variety of physiological responses. Mutation can lead to a variety of diseases, and is the focus of the current centralized study of pathology and pharmacology..GPR97 belongs to the 2 family of G protein coupled receptors, including 549 amino acid residues and a long extracellular N segment, belonging to an orphan adherent G protein coupled receptor. GPR97 may also have immune regulation. Multiple sclerosis (MS) is an autoimmune disease of the central nervous system. It is the primary disease that causes non traumatic disability in young and young people. The cause of the disease is related to a variety of factors, including heredity, environment and autoimmunity. Many studies have been made, but the effective cure method.EAE (Experimental Autoimmune Encephalomyelitis) is a common animal model for studying MS. It has many similarities with MS in pathogenesis and Clinicopathology. It is usually used to prepare EAE model by using MOG peptide to immunize mice. It has been widely used in the pathogenesis of MS and it has been widely used in the pathogenesis of MS. In the study of effective treatment methods, Dendritic cell is the most important antigen presenting cell in human body. It is an important bridge to connect inherent immunity and acquired immunity. Immature DC mainly exercises antigen phagocytosis and processing function, mature DC high expression MHC molecule and co stimulator such as CD40, CD86 and so on, and can pass through Different cytokines are secreted to induce initial Th cells to differentiate into different cell directions. A large number of studies have shown that DC cells may have a close relationship with the pathogenesis of EAE..GPR97 knockout mice are a powerful tool to study the function of the GPR97 gene. We detected the immune cells and subgroups in the splenocytes of the GPR97 gene knockout mice. The results showed that the number of T cells in the splenocytes of GPR97 knockout mice was slightly lower than that of the wild type mice, while the number of NK cells, CD4+T cells and Thl and Th17 cells were slightly higher than those of the wild type mice. The detection results of dendritic cells showed that the number of dendritic cells in the spleen of the GPR97 gene knockout mice was significantly higher than that of the wild type mice, and the number of dendritic cells in the spleen was significantly higher than that of the wild type mice. There is a higher expression of surface molecules. In order to study the role of GPR97 in the pathogenesis of EAE, we use the GPR97 gene knockout mice as the research object and use the MOG peptide immunization to establish the EAE model. By analyzing the clinical score of the infected mice, the pathological section of the spinal cord and the detection of the related immunological indexes to determine the GPR97 gene. The effect of knockout on the pathogenesis of EAE mice showed that compared with the EAE model in the wild type mice, the EAE model of GPR97 knockout mice was higher in clinical score, the inflammatory cell infiltration in the central nervous system was more serious, the number of Th1 and Th17 cells in the spleen was more, the expression level of inflammatory factors TNF- a, IL-17A and IL-6 was higher, MOG special. The number of heterosexual T cells is more, and it is preliminarily proved that the incidence of EAE in mice with GPR97 knockout is more serious. In order to further explore the possible mechanism of GPR97 affecting the pathogenesis of EAE, we use DC cells as the target cells to detect the cytokine secreting spectrum of DC cells, the expression of the surface molecules, the differentiation of Th and the antigen presentation of the GPR97 gene knockout. The effects of biological functions such as.RT-PCR and ELISA showed that the GPR97 knockout DC secreted more IL-6, TGF- beta and IL-12, and the secretion of IL-10 decreased; the GPR97 knockout DC cells mainly induced the Th cells to differentiate into Th1 and cells; The PR97 gene knockout DC cells have a stronger ability to stimulate the proliferation of MOG specific T cells. These results suggest that the GPR97 gene knockout enhances the biological function of DC cells. Therefore, the results of this study suggest that the GPR97 gene is associated with the pathogenesis of EAE mice. The possible mechanism of this study is that GPR97 based knockout enhances the immune function of DC. Objective to explore a theoretical and experimental basis for the treatment of MS with GPR97 as the target.

【學(xué)位授予單位】：華東師范大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R744.51

【共引文獻(xiàn)】

相關(guān)期刊論文 前9條

1 毛悅時(shí),呂傳真;多發(fā)性硬化的流行病學(xué)[J];國(guó)外醫(yī)學(xué).神經(jīng)病學(xué)神經(jīng)外科學(xué)分冊(cè);2004年04期

2 李莉;楊照華;王慧敏;;52例多發(fā)性硬化臨床分析[J];中國(guó)實(shí)用神經(jīng)疾病雜志;2014年19期

3 樊永平;尤昱中;陳克龍;楊濤;吳畏;;261例多發(fā)性硬化患者臨床特點(diǎn)和中醫(yī)證候分布[J];首都醫(yī)科大學(xué)學(xué)報(bào);2012年03期

4 Chisen Takeuchi;Kanato Yamagata;Takako Takemiya;;Variation in experimental autoimmune encephalomyelitis scores in a mouse model of multiple sclerosis[J];World Journal of Neurology;2013年03期

5 初曙光;李振新;馮曉源;;多發(fā)性硬化的磁共振成像診斷及研究進(jìn)展[J];中國(guó)現(xiàn)代神經(jīng)疾病雜志;2011年03期

6 曹小麗;秦嬌琴;;63例視神經(jīng)脊髓炎患者臨床特點(diǎn)及尿酸水平分析[J];現(xiàn)代預(yù)防醫(yī)學(xué);2012年15期

7 王薇;文程;陳英才;王華;;神經(jīng)生長(zhǎng)因子對(duì)EAE大鼠腦神經(jīng)干細(xì)胞增殖以及對(duì)小兒病毒性腦炎的影響研究[J];中國(guó)婦幼保健;2014年17期

8 趙志斌;;4例家族性多發(fā)性硬化病患者特點(diǎn)分析[J];中國(guó)民康醫(yī)學(xué);2015年06期

9 褚海波;王磊;徐永波;王濤;鞠玲燕;菅鳳國(guó);趙建華;;門靜脈高壓巨脾大部切除后殘脾神經(jīng)分布與定量研究[J];中國(guó)普通外科雜志;2015年09期

相關(guān)博士學(xué)位論文 前5條

1 李俊敏;MS易感基因、認(rèn)知障礙與細(xì)胞免疫治療機(jī)制的研究[D];華中科技大學(xué);2011年

2 毛悅時(shí);病毒感染與多發(fā)性硬化的臨床和實(shí)驗(yàn)室研究[D];復(fù)旦大學(xué);2006年

3 孫海波;慢丙肝抗病毒治療中依從性和療效的關(guān)系及CD24預(yù)測(cè)病毒清除的價(jià)值[D];吉林大學(xué);2014年

4 陳炳偉;Th17細(xì)胞相關(guān)炎癥因子與冠心病及動(dòng)脈硬化關(guān)系的研究[D];北京協(xié)和醫(yī)學(xué)院;2014年

5 馮博;IL1β通過(guò)上調(diào)CB1R促進(jìn)幼鼠熱驚厥后成年癲癇易感性[D];浙江大學(xué);2014年

相關(guān)碩士學(xué)位論文 前5條

1 田美娟;利比添加治療多發(fā)性硬化的前瞻開(kāi)放對(duì)照研究[D];華中科技大學(xué);2011年

2 李予巧;NMO-IgG及CD4+T輔助細(xì)胞相關(guān)細(xì)胞因子在MS及NMO中的檢測(cè)及意義[D];復(fù)旦大學(xué);2009年

3 佘曉嵐;不同劑量干擾素-β對(duì)多發(fā)性硬化療效及安全性的系統(tǒng)評(píng)價(jià)[D];山西醫(yī)科大學(xué);2012年

4 梁斐;Parkin基因多態(tài)性與急性一氧化碳中毒后遲發(fā)性腦病的關(guān)聯(lián)研究[D];新鄉(xiāng)醫(yī)學(xué)院;2012年

5 任海燕;視神經(jīng)脊髓炎疾病譜和多發(fā)性硬化的臨床研究[D];天津醫(yī)科大學(xué);2010年

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本文編號(hào)：1882078