本文選題：腎結(jié)石 + SLC26A6蛋白　； 參考：《遼寧大學(xué)》2017年碩士論文

【摘要】：腎結(jié)石是由遺傳因素和飲食環(huán)境等因素引起的一種常見的、多發(fā)的復(fù)雜疾病,具有較高的復(fù)發(fā)率,形成機(jī)制尚不明確,目前缺乏較為有效的預(yù)防和治療措施。草酸鈣結(jié)石是目前最為常見的腎結(jié)石類型,約占全部腎結(jié)石種類的80%以上。草酸是生物體中的一種常見代謝產(chǎn)物,能夠使礦質(zhì)元素的生物利用率降低,在人體中容易與鈣離子結(jié)合,形成草酸鈣,進(jìn)而導(dǎo)致腎結(jié)石疾病的發(fā)生。尿中焦磷酸是重要的結(jié)晶形成抑制物,可降低尿中鈣、磷等離子的過飽和狀態(tài),在腎小管中可結(jié)合到磷酸鈣表面,抑制結(jié)晶進(jìn)一步形成。SLC26A6蛋白是一個(gè)多功能的陰離子轉(zhuǎn)運(yùn)蛋白,尤其在轉(zhuǎn)運(yùn)草酸陰離子的過程中發(fā)揮著重要的作用。進(jìn)行性關(guān)節(jié)強(qiáng)直基因同源物ANKH(progressive ankylosis gene homolog,Ank/ANKH)是一種無機(jī)焦磷酸鹽的轉(zhuǎn)運(yùn)蛋白(inorganic pyrophosphate transport regulator),能夠在腎臟表達(dá),在集合管的腔面和基底面,向腎小管和腎間質(zhì)轉(zhuǎn)運(yùn)焦磷酸。推測這兩種蛋白可能在腎結(jié)石形成過程中發(fā)揮重要作用。本文首先以SLC26A6基因和ANKH基因?yàn)閷ο?聯(lián)合采用幾種基因致病型SNP預(yù)測生物信息學(xué)軟件,分別預(yù)測出兩種基因可能導(dǎo)致疾病的SNPs。然后以腎結(jié)石疾病患者和健康人的血液樣本進(jìn)行基因型檢測,以驗(yàn)證預(yù)測的兩種基因的致病型SNPs是否與腎結(jié)石的發(fā)病相關(guān),并研究其基因多態(tài)性與腎結(jié)石疾病發(fā)病風(fēng)險(xiǎn)的關(guān)系。我們從腎結(jié)石疾病患者和健康人的血液中提取人全基因組DNA,并以其為PCR擴(kuò)增模板,通過等位基因特異性PCR及基因測序方法對SLC26A6基因中經(jīng)生物信息學(xué)軟件預(yù)測得到的1個(gè)候選SNP以及ANKH基因中的14個(gè)可能的致病候選SNPs進(jìn)行基因定型。相關(guān)分析發(fā)現(xiàn),在SLC26A6基因中,腎結(jié)石患者nsSNP rs184187143的C等位基因率顯著高于無腎結(jié)石病史以及結(jié)石疾病家族史的健康人對照樣本(OR 6.1,95%CI 1.36-27.38,p=0.007)。而所選擇驗(yàn)證的ANKH基因中14個(gè)位點(diǎn)的SNPs與東北地區(qū)漢族草酸鈣腎結(jié)石發(fā)病的風(fēng)險(xiǎn)并無明顯關(guān)系。最后,我們利用分子動(dòng)力學(xué)方法對SLC26A6基因致病nsSNP rs184187143導(dǎo)致腎結(jié)石患病風(fēng)險(xiǎn)增加的分子機(jī)制進(jìn)行進(jìn)一步研究,結(jié)果發(fā)現(xiàn)SLC26A6基因致病nsSNP rs184187143位點(diǎn)處,等位基因G到C的變化(導(dǎo)致氨基酸從甘氨酸到精氨酸的突變)可能通過影響蛋白的二級結(jié)構(gòu),影響其草酸離子轉(zhuǎn)運(yùn)功能,從而與腎結(jié)石發(fā)病風(fēng)險(xiǎn)相關(guān)。綜上,本課題首次證明了SLC26A6基因多態(tài)性變異nsSNP rs184187143影響中國東北地區(qū)漢族草酸鈣腎結(jié)石發(fā)病風(fēng)險(xiǎn),暗示草酸轉(zhuǎn)運(yùn)過程的異�？赡茇暙I(xiàn)于腎結(jié)石的形成。這一研究成果為腎結(jié)石疾病的復(fù)雜分子機(jī)制的研究提供了新的思路。
[Abstract]:Kidney stone is a common and complex disease caused by genetic factors and dietary environment. It has a high recurrence rate and its formation mechanism is not clear. At present, it is lack of effective prevention and treatment measures.Calcium oxalate stone is the most common type of renal calculi, accounting for more than 80% of all renal calculi.Oxalic acid is a common metabolite in organism, which can reduce the bioavailability of mineral elements, bind easily with calcium ion in human body, form calcium oxalate, and lead to the occurrence of kidney stone disease.Urinary pyrophosphoric acid is an important inhibitor of crystallization formation, which can reduce the supersaturated state of calcium and phosphorus plasma in urine. It can bind to the surface of calcium phosphate in renal tubules and inhibit the further formation of crystallization. SLC26A6 protein is a multifunctional anion transporter.Especially in the process of oxalic acid anion transport plays an important role.The progressive ankylosis gene congener, ANKH(progressive ankylosis gene homologue, is an inorganic pyrophosphate transport protein, the organic pyrophosphate transport regulator, that can be expressed in the kidney and transport pyrophosphoric acid to the renal tubules and the interstitial surface in the lumen and basal plane of the collecting duct.It is speculated that these two proteins may play an important role in the formation of renal calculi.In this paper, SLC26A6 gene and ANKH gene were used to predict SLC26A6 gene and ANKH gene respectively by using several gene pathogenic SNP predictive bioinformatics software.Then genotypes were detected in the blood samples of patients with renal stone disease and healthy people to verify whether the predicted pathogenic SNPs of the two genes was related to the pathogenesis of renal calculi and to study the relationship between the gene polymorphism and the risk of renal stone disease.We extracted human genomic DNA from the blood of patients with kidney stone disease and healthy people and used it as a template for PCR amplification.Allele-specific PCR and gene sequencing were used to identify one candidate SNP and 14 possible pathogenicity candidate SNPs in SLC26A6 gene and ANKH gene predicted by bioinformatics software.Correlation analysis showed that the C allele rate of nsSNP rs184187143 in patients with renal calculi was significantly higher than that in healthy controls with no history of renal calculi and family history of calculi.However, there was no significant correlation between SNPs of 14 loci of the selected ANKH gene and the risk of calcium oxalate nephrolithiasis in Northeast China.Finally, we further studied the molecular mechanism of the increase in the risk of renal calculi caused by SLC26A6 gene pathogenic nsSNP rs184187143 by molecular dynamics. The results showed that the site of SLC26A6 gene pathogenic nsSNP rs184187143 was found.The change of allele G to C (leading to the mutation of amino acid from glycine to arginine) may affect the oxalate ion transport function of the protein by affecting its secondary structure, which may be related to the risk of renal calculi.In conclusion, this study for the first time proved that SLC26A6 gene polymorphism nsSNP rs184187143 affects the risk of calcium oxalate nephrolithiasis in Northeast China, suggesting that the abnormal oxalic acid transport process may contribute to the formation of renal calculi.This study provides a new idea for the study of complex molecular mechanism of renal stone disease.
【學(xué)位授予單位】：遼寧大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R692.4

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本文編號：1770317