Survivin和Apollon在肝细胞癌中的表达及分析研究

发布时间：2018-08-21 12:19

【摘要】：目的通过改良型间断饮用二乙基亚硝胺（diethylnitrosamine,DEN）诱导大鼠原发性肝癌模型，观察大鼠肝脏在不同诱癌阶段的病理变化，同时检测Survivin和Apollon在大鼠肝脏不同诱癌阶段的表达情况，旨在研究Survivin和Apollon在诱导的大鼠肝细胞癌发生、发展过程中的变化，探讨Survivin和Apollon作为肝癌早期诊断标志物的可行性；收集部分临床人肝癌标本及其癌旁组织、肝良性病变组织，检测Survivin和Apollon表达情况，并结合临床病理参数，探讨Survivin和Apollon在人肝癌形成过程中的意义。方法 4周龄的雌雄性近交系wista大鼠共80只，随机分为实验组（A组，n=68）和对照组（B组，n=12）。A组动物间断饮用0.01%二乙基亚硝胺(DEN)溶液，B组自由饮用灭菌自来水。实验中观察大鼠的一般情况，于实验第4周、8周、12周、16周、18周分别取A组大鼠12只，B组大鼠2-3只，称重后麻醉，开腹摘取大鼠肝脏，观察肝脏大体情况及组织病理学变化；分别采用免疫组织化学法检测Survivin和Apollon在大鼠肝癌形成不同阶段的表达情况，并将两者在各阶段表达情况进行比较分析；用免疫组织化学方法检测Survivin和Apollon蛋白在42例原发性肝癌组织及其癌旁组织、15例肝脏良性病变组织的表达情况，，实验数据结合临床病理参数进行统计学分析。结果 1.改良型间断饮用DEN成功诱导了大鼠肝癌模型，大鼠肝脏癌变大致经过肝细胞损伤期、肝硬化期和肝细胞癌变期三个阶段。至诱癌结束时实验组大鼠自然死亡4只，死亡率5.88%（4/68）。实验组第18周诱癌结束时存活的16只大鼠15只形成肝癌，诱癌率93.75%（15/16）。 2.Survivin和Apollon在正常大鼠肝脏组织不表达，伴随着肝癌的发展，Survivin和Apollon蛋白阳性表达率和表达强度依次升高，各组间阳性率比较差异有统计学意义（p0.05）。与对照组相比，Survivin和Apollon在肝硬化期、肝癌变期阳性率均上升（P0.05/3），肝癌变组与肝硬化组比较，Survivin和Apollon的阳性表达率亦出现上升(P0.05/3)。 3.Survivin蛋白在临床肝癌组、癌旁组及良性组的阳性率分别为71.43%（30/42）、40.48%(17/42)、0.00%（0/15），各组间差异有统计学意义（p0.05）。Survivin蛋白在HCC中的表达与患者性别、年龄、HBsAg、AFP、肿瘤大小无明显相关（p0.05），而与病理分级、门静脉癌栓相关（p0.05）。 4.Apollon蛋白在临床肝癌组、癌旁组及良性组的阳性率分别为66.67%（28/42)、38.09%(16/42)、0.00%（0/15），各组间差异有统计学意义（p0.05）。Apollon蛋白在HCC中的表达与患者性别、年龄、HBsAg、AFP、肿瘤大小无明显相关（p0.05），而与病理分级、门静脉癌栓相关（p0.05）。 5.两者相关性：临床人肝癌组织中Survivin和Apollon蛋白的表达呈正相关关系（r=0.409,p=0.005）。结论 1.本研究通过间断饮用DEN成功诱导了大鼠肝癌模型，该模型简单、安全有效，诱癌大鼠具有低死亡率和高诱癌率的特点。 2.Survivin和Apollon参与了大鼠肝癌的发生、发展，对于大鼠肝癌发生各个阶段均有提示作用，提示其可作为肝癌早期诊断的潜在标志物。 3.Survivin和Apollon在人肝癌组表达明显高于良性对照组，说明其表达异常和失衡与肝癌的发生、发展密切相关。Survivin和Apollon与门脉癌栓的形成及HCC的病理分级有一定的关系，提示Survivin和Apollon可作为判断肝癌侵袭和转移的指标。 4.Survivin和Apollon在人肝癌旁组织中表达与良性对照组有统计学意义，提示阳性表达的癌旁肝组织具有癌变危险性，癌旁肝组织可能是一群异常过度增生的肝细胞群，其中部分细胞有可能发展成为HCC。 5.大鼠肝癌的Survivin和Apollon蛋白表达情况和人肝癌的表达情况基本一致，该模型能较好地从分子层面模拟人类肝癌发生，为今后肝癌的研究提供了有效的手段。
[Abstract]:objective
To study the effect of diethylnitrosamine (DEN) on hepatocellular carcinoma (HCC) induced by diethylnitrosamine (DEN) in rats, the pathological changes of rat liver in different stages of carcinogenesis were observed, and the expressions of Survivin and Apollon in different stages of hepatocarcinogenesis were detected. To explore the feasibility of Survivin and Apollon as early diagnostic markers of hepatocellular carcinoma, to collect some clinical specimens of human hepatocellular carcinoma and its adjacent tissues, benign liver lesions, to detect the expression of Survivin and Apollon, and to explore the role of Survivin and Apollon in the formation of human hepatocellular carcinoma in combination with clinicopathological parameters. Significance.
Method
Eighty four-week-old male and female WISTA rats were randomly divided into experimental group (group A, n=68) and control group (group B, n=12). The rats in group A were given 0.01% diethylnitrosamine (DEN) solution intermittently and the rats in group B were given tap water freely. Two to three rats were anesthetized after weighing and abdominal liver was taken out to observe the general situation and histopathological changes of the liver. The expression of Survivin and Apollon in different stages of hepatocarcinogenesis was detected by immunohistochemical method, and the expression of Survivin and Apollon in different stages were compared and analyzed. Survivin and Apollon proteins were detected in 42 cases of primary hepatocellular carcinoma and its adjacent tissues, and 15 cases of benign liver lesions. The experimental data were analyzed statistically with clinicopathological parameters.
Result
1. Modified intermittent drinking of DEN successfully induced hepatocarcinogenesis in rats. The hepatocarcinogenesis of the rats went through three stages: hepatocyte injury stage, liver cirrhosis stage and hepatocyte carcinogenesis stage. The cancer rate was 93.75% (15/16).
2. Survivin and Apollon were not expressed in normal rat liver tissues. With the development of hepatocellular carcinoma, the positive expression rate and intensity of Survivin and Apollon protein increased in turn, and the difference was statistically significant (p0.05). The positive expression rates of Survivin and Apollon in HCC group were higher than those in cirrhosis group (P0.05/3).
3. The positive rate of Survivin protein was 71.43% (30/42), 40.48% (17/42) and 0.00% (0/15) in clinical hepatocellular carcinoma group, adjacent cancer group and benign group, respectively. The difference was statistically significant (p0.05). The expression of Survivin protein in HCC was not significantly correlated with gender, age, HBsAg, AFP, tumor size (p0.05), but with pathological grade and portal vein tumor thrombus (p0.05). P0.05).
4. The positive rate of Apollon protein was 66.67% (28/42), 38.09% (16/42) and 0.00% (0/15) in clinical hepatocellular carcinoma group, adjacent cancer group and benign group, respectively. The difference was statistically significant (p0.05). The expression of Apollon protein in HCC was not significantly correlated with gender, age, HBsAg, AFP, tumor size (p0.05), but with pathological grade and portal vein tumor thrombus (p0.05). .05).
5. The correlation between the expression of Survivin and Apollon protein was positive (r = 0.409, P = 0.005).
conclusion
1. In this study, a rat model of hepatocellular carcinoma was successfully induced by intermittent drinking of DEN. The model was simple, safe and effective, and the induced cancer rats had the characteristics of low mortality and high induced cancer rate.
2. Survivin and Apollon are involved in the occurrence and development of hepatocellular carcinoma in rats, which may be a potential marker for early diagnosis of hepatocellular carcinoma.
3. The expression of Survivin and Apollon in HCC group was significantly higher than that in benign control group, indicating that the abnormal and unbalanced expression of Survivin and Apollon was closely related to the occurrence and development of HCC.
4. The expression of Survivin and Apollon in the tissues adjacent to HCC was statistically significant with that in the benign control group, suggesting that the positive expression of Survivin and Apollon in the tissues adjacent to HCC is at risk of carcinogenesis.
5. The expression of Survivin and Apollon protein in rat hepatocellular carcinoma is basically the same as that in human hepatocellular carcinoma. The model can simulate the occurrence of human hepatocellular carcinoma from the molecular level and provide an effective means for the future study of hepatocellular carcinoma.
【学位授予单位】：广州医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R735.7

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