本文選題：原發(fā)性肝癌　切入點(diǎn)：細(xì)胞凋亡　出處：《天津醫(yī)科大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：【目的】分析原發(fā)性肝癌患者血清及病理組織中uMtCK的表達(dá)水平,初步探討uMtCK對(duì)輔助肝癌臨床診斷及預(yù)后判斷的價(jià)值;分析uMtCK對(duì)肝癌細(xì)胞增殖、凋亡、侵襲及遷移作用及機(jī)制,探討uMtCK與肝癌細(xì)胞惡性生物學(xué)特征的關(guān)系。【方法】在臨床血清中,采用ELISA方法,檢測(cè)uMtCK蛋白在慢性乙型肝炎、活動(dòng)性肝硬化和原發(fā)性肝癌患者血清中的表達(dá)水平、uMtCK蛋白在肝癌不同臨床分期患者血清中的表達(dá)水平以及肝癌患者RFA根治性治療前后血清中uMtCK蛋白的表達(dá)水平;根據(jù)受試者特征曲線得到曲線下面積(Area under the curve,AUC)探討uMtCK和AFP對(duì)輔助肝癌的診斷價(jià)值及聯(lián)合診斷價(jià)值;在臨床組織標(biāo)本中,采用免疫組織化學(xué)的方法,檢測(cè)uMtCK蛋白在慢性乙型肝炎、活動(dòng)性肝硬化和原發(fā)性肝癌不同組織中的表達(dá)水平;采用logistic回歸方法分析uMtCK蛋白表達(dá)水平與肝癌不同臨床指標(biāo)和肝癌惡性生物學(xué)特征的關(guān)系;在體外肝癌細(xì)胞株中采用Real-time PCR方法檢測(cè)uMtCK蛋白在不同肝癌細(xì)胞株中的表達(dá)水平;MTT比色法檢測(cè)uMtCK基因?qū)Ω伟┘?xì)胞的增殖作用;流式細(xì)胞術(shù)檢測(cè)uMtCK基因?qū)Ω伟┘?xì)胞凋亡的作用機(jī)制;并采用si RNA技術(shù)靶向沉默uMtCK基因表達(dá),采用Brdu細(xì)胞增殖試驗(yàn)和Transwell小室腫瘤細(xì)胞遷移與侵襲實(shí)驗(yàn)分析探討uMtCK基因表達(dá)沉默對(duì)肝癌細(xì)胞株生物學(xué)特性的影響。【結(jié)果】1、uMtCK蛋白在原發(fā)性肝癌患者組血清中的表達(dá)水平高于活動(dòng)性肝硬化組和慢性乙型肝炎組(P0.05);uMtCK蛋白在晚期肝癌亞組患者血清中的表達(dá)水平高于早期肝癌亞組(P0.05);肝癌患者血清中uMtCK蛋白濃度為10 U/L臨界點(diǎn)診斷各期肝癌組與正常組時(shí),uMtCK檢測(cè)肝癌的特異度低于AFP(69.8%vs77.4%,P0.05);AFP與uMtCK診斷肝癌的靈敏度及AUC值比較差異無統(tǒng)計(jì)學(xué)意義(P0.05);AFP與uMtCK聯(lián)合診斷肝癌的靈敏度及Youden指數(shù)高于AFP(P0.05);肝癌組患者RFA根治術(shù)治療后血清中uMtCK表達(dá)水平較術(shù)前下降(P0.05)。2、uMtCK蛋白在原發(fā)性肝癌組織中表達(dá)水平高于慢性乙型肝炎及正常肝組織(P0.05);且uMtCK蛋白表達(dá)水平與有無門靜脈受侵及腫瘤病理分級(jí)呈正相關(guān)(P0.05)。3、uMtCK基因在體外肝癌細(xì)胞株Hep G2、Hu H7中呈高表達(dá),且穩(wěn)定高表達(dá)uMtCK在Hep G2、Hu H7的肝癌細(xì)胞株中有促進(jìn)肝癌細(xì)胞增殖作用(P0.05);穩(wěn)定高表達(dá)uMtCK在Hep G2肝癌細(xì)胞株中有抑制肝癌細(xì)胞凋亡作用(P0.05)。應(yīng)用si RNA技術(shù)靶向沉默uMtCK基因表達(dá)后,肝癌細(xì)胞增殖能力顯著下降(P0.05),且肝癌細(xì)胞的遷移和侵襲能力明顯受到抑制(P0.05)。【結(jié)論】uMtCK蛋白在肝癌患者血清及病理組織中高表達(dá);uMtCK與AFP聯(lián)合檢測(cè)診斷肝癌的靈敏度及Youden指數(shù)高于AFP;肝癌Ⅲ-Ⅳ期亞組的uMtCK血清表達(dá)水平較顯著高于肝癌Ⅰ-Ⅱ期亞組,uMtCK在肝癌病理組織表達(dá)強(qiáng)度與門靜脈癌栓及病理分化水平呈正相關(guān);uMtCK在肝癌細(xì)胞株Hep G2、Hu H7中穩(wěn)定高表達(dá)可抑制肝癌細(xì)胞凋亡,在肝癌細(xì)胞的X椫場(chǎng)⑶ㄒ啤⑶窒裥隕镅形釁鹱糯俳饔謾MtCK有可能成為輔助肝癌血清學(xué)診斷標(biāo)記物,對(duì)于肝癌臨床分期及預(yù)后判斷有一定的監(jiān)測(cè)指導(dǎo)意義。
[Abstract]:[Objective] analyze the expression level of primary uMtCK in serum and pathological tissues of patients with hepatocellular carcinoma, preliminary study of uMtCK on diagnosis and prognosis of hepatocellular carcinoma clinical auxiliary value; analysis of uMtCK on hepatocellular carcinoma cell proliferation, apoptosis, invasion and migration effect and mechanism, to explore the relationship between uMtCK and malignant biological characteristics. [method] in in serum, using ELISA method to detect the expression of uMtCK in chronic hepatitis B, liver cirrhosis and primary hepatocellular carcinoma patients serum expression level of uMtCK protein expression in hepatocellular carcinoma in different clinical stages in the serum of patients with liver cancer patients, RFA expression level of uMtCK protein in serum before and after the treatment according to the subjects; the characteristics of curve area under the curve (Area under the curve, AUC) and to evaluate the value of uMtCK and AFP in diagnosis of hepatocellular carcinoma and auxiliary combined diagnosis in clinical specimens; , using immunohistochemical method, the expression of uMtCK protein in chronic hepatitis B, the expression level of liver cirrhosis and primary liver cancer in different tissues; using logistic regression methods to analyze the relationships between the expression of uMtCK protein level and liver cancer in different clinical indicators and malignant biological characteristics; the expression level of uMtCK protein was detected in different hepatocellular carcinoma cell lines using Real-time PCR method in vitro in hepatocellular carcinoma cell line; proliferation detection uMtCK colorimetry of MTT gene on hepatocellular carcinoma cells; for the mechanism of uMtCK gene were detected by flow cytometry on apoptosis of hepatocellular carcinoma cells; and using Si RNA targeted expression of uMtCK gene silencing, the analysis of the effect of uMtCK gene silence on biological the characteristics of hepatocellular carcinoma cell line Brdu cell proliferation assay and Transwell cell tumor cell migration and invasion assay. [results] 1, uMtCK protein in primary The expression level of patients with hepatocellular carcinoma in serum was higher than that of active cirrhosis group and chronic hepatitis B group (P0.05); the expression level of uMtCK protein in the subgroup of patients with advanced hepatocellular carcinoma in serum was higher than that of early cancer subgroups (P0.05); serum uMtCK protein concentration was 10 U/L critical point diagnosis of various stages of liver cancer and normal group group, uMtCK detection of liver cancer specificity is less than AFP (69.8%vs77.4%, P0.05); there was no significant difference between the sensitivity of AUC and AFP and uMtCK in the diagnosis of hepatocellular carcinoma (P0.05); the value of the sensitivity of Youden and AFP combined with uMtCK in diagnosis of hepatocellular carcinoma (P0.05) refers to the number of higher than AFP; liver cancer patients RFA radical operation in the treatment of the expression level of uMtCK decreased serum levels of.2 (P0.05), the expression of uMtCK protein in hepatocellular carcinoma tissues is higher than the level of chronic hepatitis B and normal liver tissue (P0.05); and the expression level of uMtCK protein with and without portal vein invasion and tumor disease Grading was positively correlated (P0.05).3, uMtCK gene in vitro hepatocellular carcinoma cell line Hep G2, Hu H7 showed high expression, stable and high expression of uMtCK in Hep G2, liver cancer cell line Hu in H7 can promote the proliferation of hepatocellular carcinoma cells (P0.05); overexpression of uMtCK can inhibit the apoptosis of hepatocellular carcinoma cells in Hep G2 in hepatocellular carcinoma cell line (P0.05). The application of Si RNA technology targeting uMtCK gene expression and proliferation of HCC cells decreased significantly (P0.05), and the migration and invasion of hepatocellular carcinoma cells was significantly inhibited (P0.05). [Conclusion] the high expression of uMtCK protein in serum and pathological tissues of patients with hepatocellular carcinoma; uMtCK sensitivity in combination with AFP detection in the diagnosis of HCC and Youden index was higher than that of AFP; liver cancer III - IV period subgroup uMtCK serum expression level is significantly higher than that of liver cancer stage subgroup, the intensity of uMtCK and portal vein cancer embolus and pathological differentiation expression in hepatocellular carcinoma tissues Positive correlation; uMtCK in hepatocellular carcinoma cell line Hep G2, Hu H7 in stable high expression can inhibit the apoptosis of hepatocellular carcinoma cells in hepatocellular carcinoma cells X Shan 3 at the beer field room? Pleat shape and americium meteorites???, waxy haiku? Yong man? MtCK could become the auxiliary serological diagnostic marker for HCC, there are certain the monitoring guidance for clinical liver cancer staging and prognosis.

【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R735.7

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