本文選題：SMOC2 + 原發(fā)性肝癌　； 參考：《廣東藥科大學(xué)》2017年碩士論文

【摘要】：研究背景及目的肝癌是全球范圍內(nèi)高發(fā)的一類惡性腫瘤,其中大約有90%的肝癌為原發(fā)性肝癌(HCC)。據(jù)全球癌癥調(diào)查資料顯示,肝癌發(fā)病率在所有腫瘤中排列第五,因肝癌而死亡的人數(shù)在所有惡性腫瘤中高居第三位。在中國,肝癌更是威脅我國公民生命健康的主要腫瘤之一。據(jù)中國2015年癌癥統(tǒng)計數(shù)據(jù)顯示,肝癌已成為我國實體瘤中第四位高發(fā)病率和第三位死因的癌癥。目前研究發(fā)現(xiàn)原發(fā)性肝癌的分子發(fā)病機制包括多基因及表觀遺傳的改變、染色體畸變、基因突變等,但關(guān)于HCC具體發(fā)病機制仍不明確。因此,探索與HCC發(fā)生發(fā)展相關(guān)的潛在基因并研究其在HCC發(fā)生進展過程中的作用,對于人們更好地了解HCC發(fā)生發(fā)展的分子機制具有重要意義。模塊化鈣結(jié)合蛋白2(SMOC2)屬于SPARC蛋白家族成員之一。有研究表明,SMOC2參與調(diào)節(jié)多種細(xì)胞生物學(xué)功能,如細(xì)胞周期、細(xì)胞增殖、遷移等,但關(guān)于其在腫瘤中作用的相關(guān)研究仍較少。目前,有研究利用微陣列分析技術(shù)發(fā)現(xiàn)SMOC2在多種腫瘤中呈現(xiàn)低表達,包括卵巢癌、胰腺癌等,由此可推斷SMOC2在多數(shù)腫瘤中可能為抑癌基因,但目前仍未見關(guān)于SMOC2在HCC中的表達、預(yù)防診治意義及作用的研究報道。本研究欲通過檢測SMOC2在HCC中的表達情況,結(jié)合肝癌患者病歷及隨訪資料來探討SMOC2表達與HCC患者臨床病理參數(shù)及預(yù)后的關(guān)系,并進一步探究SMOC2對HCC發(fā)展過程中的細(xì)胞生物學(xué)功能的影響,從而來明確SMOC2在HCC中的臨床診治、預(yù)防意義及生物學(xué)功能。研究方法實驗方法:采用熒光定量RT-PCR技術(shù)檢測SMOC2在肝癌組織中的轉(zhuǎn)錄水平,利用western blotting和免疫組織化學(xué)技術(shù)檢測SMOC2蛋白在肝癌標(biāo)本和肝癌細(xì)胞中的表達,探討SMOC2與肝癌患者腫瘤相關(guān)指標(biāo)的關(guān)系,并分析其表達情況與患者預(yù)后的關(guān)系。通過慢病毒轉(zhuǎn)染,構(gòu)建過表達SMOC2的HepG2和Bel-7402細(xì)胞穩(wěn)株,研究SMOC2過表達后對肝癌細(xì)胞的生長活力、運動能力等的影響。統(tǒng)計方法:肝癌組織和癌旁對照組織中的SMOC2在轉(zhuǎn)錄水平、蛋白水平上的差異性比較及相關(guān)細(xì)胞生物學(xué)功能指標(biāo)的差異性比較使用符號秩和檢驗;卡方檢驗用于分析SMOC2表達水平與HCC患者臨床病理參數(shù)間關(guān)系;生存分析用于比較SMOC2高表達組與低表達組之間總體生存期及無病進展生存期的差異;單因素、多因素Cox比例風(fēng)險回歸分析用于研究SMOC2表達及臨床病理指標(biāo)對HCC患者總體生存期及無病進展生存期的影響。研究結(jié)果1、肝癌標(biāo)本中SMOC2 mRNA轉(zhuǎn)錄水平(n=40,P0.0001)和蛋白表達水平(n=40,P=0.0101)明顯低于癌旁對照組織;較正常肝細(xì)胞系L02,SMOC2在肝癌細(xì)胞系中(Bel-7402,HepG2,QGY-7701,SK-Hep1,SMMC-7721)的表達水平下調(diào)。2、免疫組化結(jié)果顯示SMOC2免疫陽性染色主要處于胞漿中;卡方檢驗結(jié)果表明SMOC2表達水平的高低在腫瘤大小(χ~2=9.156,P=0.002)、腫瘤數(shù)量(χ~2=4.896,P=0.027)、TNM分期(χ~2=16.356,P0.001)及是否發(fā)生轉(zhuǎn)移(χ~2=6.141,P=0.013)中的分布情況存在著差異;生存分析發(fā)現(xiàn)SMOC2高表達組的總生存期(OS,P0.001)和無病進展生存期(DFS,P=0.001)明顯長于低表達組;多因素Cox回歸分析顯示SMOC2表達水平是影響患者預(yù)后的獨立因素。3、亞組分析結(jié)果顯示在AFP≤25ng/ml或400ng/ml、腫瘤長徑5cm或≥5cm及TNM分期為Ⅰ-Ⅱ的HCC患者中,SMOC2高表達組總生存率及無病進展生存率均比低表達組高(P0.05)。4、體外實驗顯示:肝癌細(xì)胞的SMOC2表達水平上調(diào)后,細(xì)胞的增殖、克隆形成以及遷移侵襲能力受到抑制,并且會阻礙肝癌細(xì)胞周期進程,誘導(dǎo)細(xì)胞周期阻滯于G0/G1期。5、小鼠成瘤實驗顯示皮下注射過表達SMOC2肝癌細(xì)胞的裸鼠腫瘤生長速度明顯低于對照組(P0.05),皮下注射過表達SMOC2肝癌細(xì)胞的裸鼠長出的腫瘤體積及重量也明顯小于對照組(P0.05)。結(jié)論與HCC發(fā)展相關(guān)的細(xì)胞生物學(xué)進程中,SMOC2發(fā)揮著抑癌基因的作用。它可作為預(yù)測HCC患者預(yù)后的有效指標(biāo),也可能作為HCC分子靶向治療的潛在靶點。此外,肝癌患者SMOC2的表達情況還可作為為患者制定三級預(yù)防方案的參考依據(jù)。
[Abstract]:Background and objective liver cancer is a worldwide type of high incidence of malignant tumors, of which about 90% of the liver cancer is primary liver cancer (HCC). According to the global cancer survey data, the incidence of liver cancer is fifth in all tumors. The number of deaths caused by liver cancer is third in all malignant tumors. In China, liver cancer is a threat. According to China's 2015 cancer statistics, liver cancer has become one of the fourth high incidence and third causes of death in China's solid tumors. The molecular pathogenesis of primary liver cancer includes polygene and epigenetic changes, chromosomal aberrations and gene mutations. The specific pathogenesis of HCC is still unclear. Therefore, exploring the potential genes associated with the development of HCC and studying its role in the progress of HCC are of great significance for people to better understand the molecular mechanism of HCC development. Modularized calcium binding protein 2 (SMOC2) is one of the members of the SPARC family. SMOC2 is involved in regulating a variety of cellular biological functions, such as cell cycle, cell proliferation, migration, etc. but there are still few related studies on its role in the tumor. At present, microarray analysis techniques have been used to detect the low expression of SMOC2 in a variety of tumors, including nesting and pancreatic cancer, and thus infer that SMOC2 is in most of the tumors. It may be a tumor suppressor gene, but there is still no study on the expression of SMOC2 in HCC, the significance and the role of prevention and treatment. This study intends to explore the relationship between the expression of SMOC2 in HCC, the case history and the follow-up data of the patients with liver cancer, to explore the relationship between the expression of SMOC2 and the clinical pathophysiological parameters and prognosis of the patients with HCC, and to further explore the SMOC2 The effect of HCC on the cell biological function in the process of development, so as to clarify the clinical diagnosis, the preventive significance and the biological function of SMOC2 in HCC. The method of research is to detect the transcriptional level of SMOC2 in the liver cancer tissues by fluorescence quantitative RT-PCR technique, and to detect SMOC2 protein by Western blotting and immunohistochemical technique. The relationship between SMOC2 and the tumor related indexes of HCC patients and the relationship between the expression of HCC and the prognosis of the patients were analyzed. The expression of HepG2 and Bel-7402 cells expressing SMOC2 were constructed by lentivirus transfection. The effects of SMOC2 overexpression on the growth activity and exercise ability of hepatoma cells were studied. Statistical methods: the difference in the transcriptional level, the protein level and the related cell biological function indexes of the SMOC2 in the liver cancer tissue and the paracancerous control tissue were compared with the sign rank sum test, and the chi square test was used to analyze the relationship between the SMOC2 expression level and the clinical pathophysiological parameters of the HCC patients; the survival analysis was used to compare the high SMOC2 level. The difference between the overall life period and the progression free survival period between the expression group and the low expression group; the single factor, multi factor Cox proportional risk regression analysis was used to study the effect of SMOC2 expression and clinicopathological indexes on the overall survival and the progression free survival of HCC patients. Results 1, the SMOC2 mRNA transcriptional level (n=40, P0.0001) in the specimens of liver cancer. The expression level of protein (n=40, P=0.0101) was significantly lower than that of the paracancerous control tissue, and the expression level of SMOC2 in the hepatocellular carcinoma cell line (Bel-7402, HepG2, QGY-7701, SK-Hep1, SMMC-7721) was down regulated by L02, and the immunohistochemical results showed that SMOC2 immunoreactive staining was mainly in the cytoplasm. The chi square test showed that the level of SMOC2 expression was high. There were differences in the tumor size (chi ~2=9.156, P=0.002), the number of tumor (x ~2=4.896, P=0.027), TNM staging (chi ~2=16.356, P0.001) and the occurrence of metastasis (chi ~2=6.141, P=0.013). The survival analysis found that the total survival time (OS, P0.001) and the progression free survival period of the high expression group of SMOC2 were significantly longer than those of low expression groups. Multiple factor Cox regression analysis showed that the expression of SMOC2 was an independent factor affecting the prognosis of the patients.3. The results of subgroup analysis showed that in AFP < 25ng/ml or 400ng/ml, the total survival rate and the progression free survival rate of SMOC2 high expression group were higher than those of low expression group (P0.05) in the AFP < 25ng/ml or 400ng/ml. The proliferation, clone formation and migration and invasion ability of hepatoma cells were inhibited, and the cell cycle progression was inhibited and the cell cycle was blocked at G0/G1.5. The tumor growth rate of nude mice subcutaneously expressed by subcutaneous injection of SMOC2 liver cancer cells was significantly lower than that of the control group, after the SMOC2 expression level of hepatoma cells was up-regulated. (P0.05) the tumor size and weight of the nude mice expressed by subcutaneous injection of SMOC2 hepatoma cells were also significantly smaller than those of the control group (P0.05). Conclusion SMOC2 plays a role in the tumor suppressor gene in the process of cell biology related to the development of HCC. It can be used as an effective indicator for predicting the prognosis of HCC patients, and may also be the potential for targeting therapy of HCC molecules. In addition, the expression of SMOC2 in patients with liver cancer can also be used as a reference for patients to develop a three level prevention plan.

【學(xué)位授予單位】：廣東藥科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R735.7

【參考文獻】

相關(guān)期刊論文 前2條

1 Sonia Pascual;Iván Herrera;Javier Irurzun;;New advances in hepatocellular carcinoma[J];World Journal of Hepatology;2016年09期

2 Bruna Scaggiante;Maryam Kazemi;Gabriele Pozzato;Barbara Dapas;Rosella Farra;Mario Grassi;Fabrizio Zanconati;Gabriele Grassi;;Novel hepatocellular carcinoma molecules with prognostic and therapeutic potentials[J];World Journal of Gastroenterology;2014年05期

，

本文編號：1806188