發(fā)布時(shí)間：2018-01-29 04:09

  本文關(guān)鍵詞： Holliday 交叉識(shí)別蛋白 HJURP 肝細(xì)胞癌　出處：《大連醫(yī)科大學(xué)》2017年碩士論文　論文類(lèi)型：學(xué)位論文

【摘要】：目的:檢測(cè)肝細(xì)胞癌組織中 HJURP(Holliday Junction Recognition Protein,Holliday交叉識(shí)別蛋白)的表達(dá)情況,探究其表達(dá)水平與預(yù)后的關(guān)系。方法:通過(guò)免疫組化、Westernblot、逆轉(zhuǎn)錄聚合酶鏈反應(yīng)和實(shí)時(shí)聚合酶鏈反應(yīng)等技術(shù)從蛋白和RNA水平上檢測(cè)HJURP的表達(dá)水平,運(yùn)用SPSS21.0(美國(guó))軟件進(jìn)行統(tǒng)計(jì)學(xué)分析。定性資料使用卡方檢驗(yàn)。定量資料的每組數(shù)據(jù)需先進(jìn)行正態(tài)檢驗(yàn),符合正態(tài)分布的使用T檢驗(yàn)或校正T檢驗(yàn),不符合正態(tài)分布的使用非參數(shù)檢驗(yàn)。先比較cancer組和paracancer組。再將cancer組的患者分別按照性別、年齡、是否有飲酒史、是否有肝炎或肝硬化、臨床分期、肝功能Child分級(jí)分為亞組。再對(duì)各個(gè)亞組的免疫組化、Westernblot、逆轉(zhuǎn)錄聚合酶鏈反應(yīng)和實(shí)時(shí)聚合酶鏈反應(yīng)進(jìn)行分析。通過(guò)電話(huà)隨訪(fǎng)的到每位患者的無(wú)病生存期、總生存期并進(jìn)行生存分析。結(jié)果:(1)PCR結(jié)果分析發(fā)現(xiàn)cancer組和paracancer組的Ct值分別是25.9652 ± 3.4866 和 31.1120± 7.3778。兩組的 ΔCt 值分別是 4.069821 ±4.6855 和8.7879±7.07246。兩組的ΔΔCt值分別是 8.4491±4.5554和 13.0779±7.0725。根據(jù)公式計(jì)算兩組的相對(duì)定量值分別是0.8774±4.0691和0.6674±2.4538,P0.001。PCR結(jié)果進(jìn)行亞組分析發(fā)現(xiàn)HJURP的表達(dá)和患者的性別、年齡、是否有肝炎或肝硬化病史、是否有飲酒史并無(wú)關(guān)系(P0.05)。但是根據(jù)臨床分期將患者分為Ⅰ期、Ⅱ期、Ⅲ期組,分別有14人,17人,9人,三組的Ct值分別是27.5474±1.3655、26.6135 ± 1.0607 和 19.1424±1.3527,P1-2=0.5760、P1-30.001、P2-30.001。三組的 ΔCt 值分別是 7.6850±0.3179、4.0306±0.3517 和-5.3967±0.7401,P1-20.001、P1-30.001、P2-30.001。三組的ΔΔCt值分別是11.8607±0.1803、9.4389±0.1750和 6.8622±0.5081,P1-20.001、P1-30.001、P2-30.001。三組的相對(duì)定量值分別是 0.0003±0.0001、0.0024±0.0008 和0.6321 ±0.2679,P1-2=0.998、P1-30.001、P2-30.001。根據(jù)患者肝功能 Child-Pugh分級(jí)將患者分為A級(jí)組、B級(jí)組、C級(jí)組,分別有11人,22人,7人,三組的Ct 值分別是 26.6322±1.6156、26.6855± 1.0166 和 18.6203±1.3778,P1-2=0.9760、P1-3=0.001、P2-30.001。三組的ΔCt 值分別是 8.1055±0.2907、3.6986±0.5547和-6.1414±0.6449,P1-20.001、P1-30.001、P2-30.001。三組的ΔΔCt 值分別是 12.0627±0.1831、9.5259±0.266 和 6.85728±0.6159,P1-20.001、P1-30.001、P2-30.001。三組的相對(duì)定量值分別是 0.0003±0.0001、0.0020±0.0006和0.8125±0.3141,P1-2=0.989、P1-30.001、P2-30.001。(2)WESTERN-BLOT分析結(jié)果顯示cancer組和paracancer組的灰度值分別是 1.2109±0.0660 和 0.7368±0.7503,P0.001。對(duì)腫瘤組患者的 WESTERN-BLOT亞組分析發(fā)現(xiàn)HJURP在蛋白水平上和性別、年齡、是否有肝炎或肝硬化病史、是否有飲酒史無(wú)關(guān)。和臨床分期(0.2645±0.1024、1.1362±0.1084、1.6556±0.1020,P1-20.010,P2-3=0.04,P11-30.01)、Child 分級(jí)(1.3286±0.0388、1.3253±0.0567、1.5776±0.1417,P1-2=0.973,P2-3=0.031,P1-3=0.055)。(3)免疫組化結(jié)果顯示cancer組的陰性、弱陽(yáng)性、中陽(yáng)性、強(qiáng)陽(yáng)性分別是7.5%、22.5%、27.5%、42.5%;paracancer組的陰性、弱陽(yáng)性、中陽(yáng)性、強(qiáng)陽(yáng)性分別是27.5%、27.5%、42.25%、2.75%(P0.001)。免疫組化的亞組分析發(fā)現(xiàn)HJURP在腫瘤組織中的表達(dá)水平和患者的性別(P=0.875)、年齡(P=0.092)、是否有飲酒史(P=0.461)、是否有肝炎或者肝硬化病史將(P=0.911)無(wú)關(guān)。但是和臨床分期(P0.001)和肝功能Child分級(jí)(P=0.026)密切相關(guān)。(4)生存分析顯示低表達(dá)組和高表達(dá)組的平均無(wú)進(jìn)展生存期分別是16.3740±1.1140月,14.2830±0.6210月,P=0.0400。但是低表達(dá)組和高表達(dá)組的平均總生存期差異不明顯。結(jié)論:(1)無(wú)論在RNA水平還是在蛋白質(zhì)水平上,腫瘤組織中的HJURP都要明顯高于癌旁組織。HJURP的表達(dá)水平和臨床分期和Child分級(jí)相關(guān),和性別、年齡、是否有飲酒史、是否有肝炎肝硬化史無(wú)關(guān)。(2)HJURP和患者的無(wú)病生存期相關(guān),但和總生存期關(guān)系不確定。
[Abstract]:Objective: to detect HJURP in hepatocellular carcinoma (Holliday Junction Recognition Protein, Holliday protein expression recognition), and explore the relationship between expression level and prognosis. Methods: by immunohistochemistry, Westernblot, reverse transcriptase polymerase chain reaction and real-time polymerase chain reaction technology from the detection of the expression levels of HJURP protein and RNA levels the use of SPSS21.0 (America) software was used for statistical analysis. The qualitative data using the chi square test. Each set of data for quantitative data must first carry on the normality test, with normal distribution using T test or calibration T test, is not in accordance with the normal distribution of the non parametric test. Comparing the cancer group and paracancer group. The patients in the cancer group respectively according to gender, age, whether there is a history of drinking, whether hepatitis or cirrhosis, clinical stage, Child classification of liver function is divided into sub groups. The immune sub group Group, Westernblot, reverse transcriptase polymerase chain reaction and real-time polymerase chain reaction analysis. Through telephone follow-up to the disease-free survival of each patient, survival analysis and overall survival. Results: (1) the results of PCR analysis showed that cancer and paracancer groups Ct value respectively is 25.9652 + 3.4866 and 31.1120 + 7.3778. two group Ct values were 4.069821 + 4.6855 and 8.7879 + two 7.07246. group of delta delta Ct = 8.4491 + 4.5554 and 13.0779 + 7.0725. according to the formula of relative quantitative value of two groups were 0.8774 + 4.0691 and 0.6674 + 2.4538, P0.001.PCR results subgroup analysis showed that the expression of HJURP and the patients gender, age, whether there is a history of hepatitis or cirrhosis, if there is no relationship between drinking history (P0.05). But according to the clinical stage of the patients were divided into phase I, II, III group, there were 14 people, 17 people, 9 people, three group of Ct = 27.5474 + 1.3655,26.6135 + 1.0607 and 19.1424 + 1.3527, P1-2=0.5760, P1-30.001, P2-30.001. three group Ct values were 7.6850 + 0.3179,4.0306 + 0.3517 and -5.3967 + 0.7401, P1-20.001, P1-30.001, P2-30.001. three groups of delta delta Ct = 11.8607 + 0.1803,9.4389 + 0.1750 and 6.8622 + 0.5081, P1-20.001, P1-30.001. The relative quantitative P2-30.001. value of three groups were 0.0003 + 0.0001,0.0024 + 0.0008 and 0.6321 + 0.2679, P1-2=0.998, P1-30.001, P2-30.001. according to the Child-Pugh classification of liver function were divided into group A, B group, C group, there were 11 people, 22 people, 7 people, three group of Ct = 26.6322 + 1.6156,26.6855 + 1.0166 and 18.6203 + 1.3778, P1-2=0.9760, P1-3=0.001, P2-30.001. three group Ct values were 8.1055 + 0.2907,3.6986 + 0.5547 and -6.1414 + 0.6449, P1-20.001, P1-30.001, P2-30.001. three The delta delta Ct = 12.0627 + 0.1831,9.5259 + 0.266 and 6.85728 + 0.6159, P1-20.001, P1-30.001, P2-30.001. relative quantitative value of three groups were 0.0003 + 0.0001,0.0020 + 0.0006 and 0.8125 + 0.3141, P1-2=0.989, P1-30.001, P2-30.001. (2) WESTERN-BLOT analysis. The results showed that the gray scale of cancer group and paracancer group respectively 1.2109 + 0.0660 and 0.7368 + 0.7503, P0.001. of tumor patients of WESTERN-BLOT subgroup analysis found that HJURP at the protein level and gender, age, whether there is a history of hepatitis or cirrhosis, whether there is a history of drinking. And the clinical stage (0.2645 + 0.1024,1.1362 + 0.1084,1.6556 + 0.1020, P1-20.010, P2-3=0.04, P11-30.01, Child (grade) 1.3286 + 0.0388,1.3253 + 0.0567,1.5776 + 0.1417, P1-2=0.973, P2-3=0.031, P1-3=0.055). (3) immunohistochemistry results showed that cancer were negative, weak positive, positive and strong positive Were 7.5%, 22.5%, 27.5%, 42.5%; paracancer were negative, weak positive, positive and strong positive were 27.5%, 27.5%, 42.25%, 2.75% (P0.001). Immunohistochemical subgroup analysis showed that the expression level of HJURP in tumor tissues and patients' sex, age (P=0.875) (P=0.092) is there, drinking history (P=0.461), whether there is hepatitis or liver cirrhosis history (P=0.911). But the independent and clinical stage (P0.001) and Child classification of liver function (P=0.026) is closely related to (4). Survival analysis showed that low expression group and high expression group average progression free survival was 16.3740 + 1.1140 in August, 14.2830 + 0.6210 months, but the P=0.0400. low expression group and high expression group average overall survival was not significantly different. Conclusion: (1) both at the RNA level or at the protein level, the expression level and clinical tumor tissues are significantly higher in HJURP tissue adjacent to cancer.HJURP staging and Ch ILD grading correlation was not related to sex, age, drinking history, and history of hepatitis cirrhosis. (2) HJURP was associated with disease-free survival but not overall survival.

【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R735.7

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