本文選題：長鏈非編碼　切入點：RNA　出處：《鄭州大學》2017年碩士論文　論文類型：學位論文

【摘要】：胃癌在全世界范圍是嚴重危害人類健康的常見的惡性腫瘤之一,發(fā)病率占全部惡性腫瘤的第4位,占消化道惡性腫瘤的首位。據(jù)統(tǒng)計,70%的胃癌病例發(fā)生在發(fā)展中國家,其中中國人群病例占據(jù)將近45%,五年生存率仍不容樂觀,病人生活質(zhì)量較差及家庭社會經(jīng)濟負擔較重,胃癌的發(fā)病機制的深入探索及研究仍是亟待解決的。隨著胃癌病因?qū)W以及胃癌發(fā)生、發(fā)展機制的逐步深入研究,我們發(fā)現(xiàn)基因與環(huán)境因素均起著至關重要的作用。隨著研究的不斷深入,長鏈非編碼RNA(long noncoding RNAs,lnc RNAs)在癌癥的發(fā)生發(fā)展機制中有著重要的調(diào)控作用,越來越多的lnc RNA與胃癌存在密切的關聯(lián)。人類結腸癌相關轉(zhuǎn)移因子-1(colon cancer associated transcript-1,CCAT1)在胃癌組織中高表達,且與腫瘤的侵襲、轉(zhuǎn)移及預后密切相關。lnc RNA遺傳多態(tài)性在胃癌易感性機制研究提供重要突破。目的研究CCAT1單核苷酸多態(tài)性(SNPs)(rs77628730,rs6989575,rs7816475以及rs6470502)與胃癌易感性的關系,并進一步探索lnc RNA CCAT1 SNPs與環(huán)境因素之間是否存在交互作用,對存在關聯(lián)的位點進行初步功能實驗驗證,為發(fā)現(xiàn)新的胃癌遺傳易感位點提供理論支持,為胃癌的早發(fā)現(xiàn)以及早期診斷提供可靠的分子標志物。方法查閱國內(nèi)外文獻數(shù)據(jù)庫以及l(fā)nc RNA數(shù)據(jù)庫,運用生物信息學網(wǎng)站及軟件篩選CCAT1上的標簽SNPs。該研究采用病例-對照方法,選取來源于醫(yī)院的460例經(jīng)病理學診斷確診的胃癌患者作為病例組,同時將性別及年齡按照頻數(shù)匹配選取來源于社區(qū)的正常對照460例。對rs77628730以及rs6470502兩SNPs位點運用聚合酶鏈式反應-限制性片段長度分析法(PCR-RFLP)對進行基因分型,rs6989575以及rs7816475采用創(chuàng)造性酶切位點分析法(CRS-PCR)。通過SPSS21.0軟件對病例組與對照組的一般特征、四個位點不同基因型以及等位基因的分布頻率之間的差異采用t檢驗以及χ2檢驗;通過Hardy-Weinberg在線分析軟件對對照組基因型頻率進行Hardy-Weinberg平衡分析;單體型分析通過SHEsis在線軟件進行;通過MDR2.0軟件對基因-環(huán)境交互作用進行分析。對正常人群中有意義位點CCAT1 rs6470502不同基因型的血清中CCAT1mRNA表達量的差異通過q RT-PCR檢測,進一步驗證CCAT1 rs6470502遺傳變異是否影響CCAT1 mRNA的表達量。進一步探討其在胃癌發(fā)生發(fā)展過程中的調(diào)控機制。結果CCAT1 rs6470502位點SNP與胃癌密切相關,與CC純合基因型相比,TT純合基因型(OR調(diào)整:1.68;95%CI:1.01-2.79)在調(diào)整吸煙,飲酒及家族史這些混雜因素后能顯著增加胃癌的發(fā)病風險;相較于C等位基因,T等位基因與胃癌的風險增加顯著相關(OR:1.94;95%CI:1.53-2.46)。相較于A等位基因而言,rs7816475位點G等位基因與胃癌的風險降低顯著相關(OR:0.78;95%CI:0.62-0.99)。按年齡、性別、吸煙史、飲酒史及胃癌家族史進行分層分析,多因素logistic回歸分析結果顯示,rs6470502位點在大于50歲年齡段人群中(OR=1.69,95%CI=1.01-2.98)以及吸煙者(OR=1.95,95%CI=1.41-2.89)中患胃癌的發(fā)病風險有不同程度的增加;rs7816475位點在胃癌家族史者(OR=1.74,95%CI=1.30-2.87)中患胃癌風險均呈現(xiàn)不同程度的增加。q RT-PCR結果顯示,CCAT1 rs6470502位點TT(2.08±0.55)、TC(1.70±0.38)和TC+TT(1.95±0.40)基因型中CCAT1 mRNA的表達量均高于rs6470502CC(1.16±0.40)基因型,差異均具有統(tǒng)計學意義(P0.05)�；�-環(huán)境因素交互作用分析結果顯示攜帶rs77628730 A等位基因,rs6989575 T等位基因以及rs6470502 T等位基因且具有吸煙史人群患胃癌的風險顯著增加(OR:3.83;95%CI:2.96-4.96)。結論1.CCAT1 rs6470502突變等位基因T以及TT基因型可顯著增加胃癌發(fā)病風險,rs7816475突變等位基因G可降低胃癌的發(fā)病風險。2.CCAT1 rs6470502 CT+TT基因型與年齡50歲相關以及吸煙者存在相關,rs7816475 AG+GG基因型與腫瘤家族史之間存在關聯(lián)。3.CCAT1基因單體型A_(rs77628730)C_(rs6989575)G_(rs7816475)T_(rs6470502)、T_(rs77628730)C_(rs6989575)G_(rs7816475)C_(rs6470502)、T_(rs77628730)T_(rs6989575)A_(rs7816475)C_(rs6470502)可以降低胃癌發(fā)病風險,單體型A_(rs77628730)T_(rs6989575)G_(rs7816475)T_(rs6470502)、T_(rs77628730)C_(rs6989575)A_(rs7816475)T_(rs6470502)可增加胃癌發(fā)病風險。4.CCAT_(1 rs6470502)基因突變可能調(diào)節(jié)CCAT1 mRNA表達量。5.吸煙史、rs77628730 A等位基因、rs6989575 T等位基因以及rs6470502 T等位基因在胃癌發(fā)生中存在交互作用。
[Abstract]:Gastric cancer is one of the serious harm to human health of the common malignant tumors in the world, the incidence rate of malignant tumor accounted for fourth, accounting for malignant tumors of the digestive tract in the first place. According to statistics, 70% gastric cancer cases occur in developing countries, which China cases occupy nearly 45% people, the five year survival rate is still not optimistic, patient life the poor quality of family and social economic burden is heavy, still further explore and study the pathogenesis of gastric cancer to be solved. With gastric cancer etiology and pathogenesis of gastric cancer, through in-depth study on development mechanism, which plays a vital role in our discovery of genetic and environmental factors. With the deepening of the study of long chain non encoding (RNA long noncoding RNAs, LNC RNAs) plays an important role in the regulation of the mechanism of the occurrence and development of cancer, gastric cancer and LNC RNA more and more closely related to human colon cancer. Related factor -1 (colon cancer associated transfer transcript-1, CCAT1) high expression in gastric cancer tissue, and tumor invasion, metastasis and prognosis of.Lnc is closely related to the genetic polymorphism of RNA provides an important breakthrough in the study of susceptibility to gastric cancer. Objective to study the mechanism of CCAT1 single nucleotide polymorphism (SNPs) (rs77628730, rs6989575, rs7816475 and rs6470502) relationship susceptibility to gastric cancer, and to further explore the interaction between LNC RNA CCAT1 SNPs and environmental factors, the preliminary function of experimental verification of the associated site, to discover new genetic susceptibility loci for gastric cancer and provide theoretical support to provide reliable molecular markers for early detection and early diagnosis of gastric cancer. Methods at home and abroad the literature database and LNC RNA database, using bioinformatics software and web site screening by case - control the label on the CCAT1 SNPs. of the study Methods selected from hospital in 460 cases confirmed by pathological diagnosis of gastric cancer patients as case group, while gender and age in normal control, frequency selection of sources in the community. In 460 cases of rs77628730 and rs6470502 two SNPs loci by analysis of polymerase chain reaction restriction fragment length (PCR-RFLP) of the gene type rs6989575 and rs7816475 by enzyme digestion sites creative analysis method (CRS-PCR). Based on the SPSS21.0 software, the case group and the control group the general characteristics, the difference between the distribution frequency of four loci and alleles by t test and 2 test; analysis software of genotypic frequency in control group was Hardy-Weinberg balance analysis online through Hardy-Weinberg; haplotype analysis was performed by SHEsis online software; gene environment interaction was analyzed by MDR2.0 software. The difference of expression level by Q RT-PCR to detect serum CCAT1mRNA significant loci CCAT1 rs6470502 genotypes in normal population, further verify the genetic variation of rs6470502 CCAT1 influence expression of CCAT1 mRNA. To further explore the process in the development of gastric cancer in the control mechanism. The results are closely related to the CCAT1 site of rs6470502 SNP and gastric cancer, compared with CC homozygous genotype, TT homozygous genotype (OR: 1.68; 95%CI:1.01-2.79) after adjusting for smoking, drinking and family history of these confounding factors could significantly increase the risk of gastric cancer; compared to the C allele, T allele increased the risk of gastric cancer and significantly correlated (OR:1.94; 95%CI:1.53-2.46) compared. In the A allele, rs7816475 risk allele G and gastric cancer decreased significantly correlated (OR:0.78; 95%CI:0.62-0.99). According to age, gender, smoking history, drinking history and familial gastric cancer The history of stratified analysis, multivariate logistic regression analysis showed that the rs6470502 locus in more than 50 years of age (OR=1.69,95%CI=1.01-2.98) and smokers (OR=1.95,95%CI=1.41-2.89) risk in gastric cancer have different degrees of increase; rs7816475 in the family history of gastric cancer (OR=1.74,95%CI=1.30-2.87) in gastric cancer risk were increased in different degrees.Q RT-PCR results showed that the CCAT1 rs6470502 site of TT (2.08 + 0.55), TC (1.70 + 0.38) and TC+TT (1.95 + 0.40) gene expression amount of CCAT1 in type mRNA was higher than that of rs6470502CC (1.16 + 0.40) genotypes, the differences were statistically significant (P0.05). The gene environment interaction analysis display rs77628730 carrying A allele rs6989575, T allele and rs6470502 T allele and has a history of smoking people suffering from gastric cancer risk increased significantly (OR:3.83; 95%CI:2.96- 4.96). Conclusion 1.CCAT1 rs6470502 mutant T allele and TT genotype significantly increased the risk of gastric cancer, rs7816475 mutant allele G can reduce the risk of.2.CCAT1 rs6470502 CT+TT genotype and age of gastric cancer at the age of 50 and the related association of smokers, the haplotypes of.3.CCAT1 gene between A_ rs7816475 genotype and AG+GG family history the tumor (rs77628730) C_ (rs6989575) G_ (rs7816475) T_ (rs6470502), T_ (rs77628730) C_ (rs6989575) G_ (rs7816475) C_ (rs6470502), T_ (rs77628730) T_ (rs6989575) A_ (rs7816475) C_ (rs6470502) can reduce the risk of gastric cancer, haplotype A_ (rs77628730) (T_ rs6989575) G_ (rs7816475) T_ (rs6470502), T_ (rs77628730) C_ (rs6989575) A_ (rs7816475) T_ (rs6470502) can increase the risk of gastric cancer (.4.CCAT_ rs6470502 1) gene mutation may regulate the expression of mRNA CCAT1.5. rs77628730 A smoking history. The allele, rs6989575 T allele and rs6470502 T allele have interaction in the occurrence of gastric cancer.

【學位授予單位】：鄭州大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R735.2

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