【摘要】：【研究背景】阿爾茨海默病(Alzheimer's disease, AD)是一種主要在老年期發(fā)生的以進行性癡呆為主要特征的神經(jīng)元退行性疾病。AD的病因尚未明確，眾多研究表明其與腦內(nèi)慢性炎癥反應(yīng)相關(guān)。IL18作為一種炎癥細胞因子，近年越來越多被證實與AD發(fā)病相關(guān)。IL18基因調(diào)控區(qū)存在－607C/A（RS1946518）、－137G/C（RS187238）等SNPs，已有文獻報道多態(tài)性與老年性癡呆發(fā)病間的關(guān)系，但未見這兩處基因多態(tài)性對IL18基因轉(zhuǎn)錄活性影響的報道。 【研究目的】本課題通過分子生物學(xué)技術(shù)，建立IL18－607C/A、IL18－137G/C野生型和突變等位基因的PGL4.10重組試驗質(zhì)粒，在細胞水平研究－607C/A突變、－137G/C突變對IL18基因轉(zhuǎn)錄調(diào)控的影響。 【方法】選取雙熒光報告基因系統(tǒng)（pGL4.10（luc2) Basic及pRL-TK)作為報告基因載體，用基因重組和定點突變技術(shù)構(gòu)建含不同SNP型的IL18啟動子報告基因質(zhì)粒，轉(zhuǎn)染人胚胎腎細胞系293細胞（HEK293)，計算不同SNP各啟動子的相對熒光素酶活性（relative luciferase activity，，RLA) 【結(jié)果】所構(gòu)建的各SNP型質(zhì)粒均具有明確的啟動子活性；突變質(zhì)粒-607A/-137C、-607C/-137G及-607C/-137C與野生質(zhì)粒-607A/-137G相比，轉(zhuǎn)錄活性均有顯著性差異，（P0.01）；而單位點突變質(zhì)粒-607A/-137C、-607C/-137G與兩位點突變質(zhì)粒-607C/-137C轉(zhuǎn)錄活性相比均無差異性(P0.05)。 【結(jié)論】-607、-137位點堿基的突變均可影響IL18基因的轉(zhuǎn)錄，-607A/-137C、-607C/-137G及-607C/-137C均上調(diào)了IL18基因的轉(zhuǎn)錄活性，致IL18蛋白表達水平增加，促進神經(jīng)炎癥的病理過程，可能導(dǎo)致AD的發(fā)生。
[Abstract]:[background] Alzheimer's disease (Alzheimer's disease, AD) is a neuronal degenerative disease characterized mainly by progressive dementia in the elderly. The etiology of AD is not clear. IL18, as an inflammatory cytokine, has been more and more associated with the pathogenesis of AD in recent years. SNPs, exists in the regulatory region of IL18 gene such as 607C/A (RS1946518),-137G/C (RS187238) and so on. The relationship between polymorphism and Alzheimer's disease has been reported, but there is no report on the effect of these two gene polymorphisms on the transcriptional activity of IL18 gene. [objective] by using molecular biology technique, the PGL4.10 recombinant plasmid of IL18-607C/A,IL18-137G/C wild-type and mutant alleles was established, and the 607C/A mutation was studied at the cell level. The effect of 137G/C mutation on the transcriptional regulation of IL18 gene. [methods] double fluorescent reporter gene systems (pGL4.10 (luc2) Basic and pRL-TK) were selected as reporter gene vectors. The IL18 promoter reporter gene plasmids containing different SNP types were constructed by gene recombination and site-directed mutagenesis. Transfected into human embryonic kidney cell line 293 (HEK293), the relative luciferase activity (relative luciferase activity,RLA) of different SNP promoters was calculated. [results] all the SNP type plasmids constructed showed definite promoter activity. The transcriptional activities of mutant plasmids -607A / -137C / -607C / -137G and-607C/-137C were significantly different from those of wild plasmids-607A/-137G (P0.01). However, the transcriptional activity of the unit point mutant plasmide-607A / -137C / -607C / -137G was not different from that of the two locus mutant plasmids-607C/-137C (P0.05). [conclusion] the mutation at -607C ~ 137 can affect the transcription of IL18 gene. -607A / -137C / -137C / -137G and-607C/-137C can up-regulate the transcriptional activity of IL18 gene and increase the expression of IL18 protein. Promoting the pathological process of neuroinflammation may lead to the occurrence of AD.
【學(xué)位授予單位】：廣州醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R749.16

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