【作者】 高建鹏；

【导师】 郭永章； 李立；

【作者基本信息】 昆明医学院 ， 普通外科学， 2005， 博士

【摘要】 提高人类原发性肝癌的疗效仍是医学界面临的难题。本研究利用生物治疗探索干预人原发性肝癌复发的治疗方案。基于细胞分子应答技术研究生物联合干预对肝癌细胞分化、细胞周期调控,癌组织血管抑制和免疫调控因子及基因表达谱的变化。应用基因芯片检测了人肝癌细胞BEL-7402细胞株的生物信息,体内、外分别使用生长抑素8肽、生长抑素8肽联合IFN—α2b干扰素调控肝癌细胞的分子应答机制。在不同时间,实验条件下,通过相差显微镜,苔盼蓝拒染法,流式细胞仪,基因芯片等检测手段。我们发现BEL-7402细胞与正常肝细胞在基因表达谱上有明显差异,在BEL-7402细胞系阳性表达的基因有79个。与正常人肝细胞表达谱有表达差异的基因有31个,上调24个,下调7个。生长抑素8肽体外实验发现对人肝癌细胞的抗增殖作用是通过影响细胞分化、细胞周期分布实现。表达差异的基因主要与G、S周期调控有关。细胞凋亡不是主要的作用机制。裸小鼠背部皮下注射BEL-7402细胞悬液,建立人原发性肝癌裸小鼠皮下移植模型。根据不同分组皮下注射生长抑素和生长抑素联合干扰素进行干预。通过观察皮下肿瘤生长情况,病理学检查,原位末端标记流式细胞仪检测细胞凋亡,免疫组化,微血管密度检测,基因芯片检测基因表达谱的变化。研究结果显示单独使用生长抑素对抑制人肝癌细胞移植瘤的增殖有作用,而联合使用生长抑素加干扰素则表现出更明显的肿瘤抑制效应。我们认为生长抑素抗人肝癌细胞移植瘤的作用机制是影响细胞的周期分布、细胞分化以及干预肿瘤血管生成。生长抑素联合干扰素使用对肿瘤细胞影响在于影响细胞周期分布,抑制了肿瘤新生血管的生成并且有促进瘤细胞凋亡的作用。基因表达谱显示23个基因有表更多还原

【Abstract】 To investigate molecular characters of BEL-7402 , it were studied under cDNA microarray. Then to observe the effects of Somatostatin with Interferon-alpha inhibited proliferation of BEL-7402 hepatocellular carcinoma(HCC) cell line and growth of HCC xenografts in nude mice. Finally , to evaluate security of administration of Somatostatin in HCC patients after resection of liver. We compared gene expression alteration between BEL-7402 cell line and normal liver tissue by cDNA microarray. Among 866 cancer related genes, there were 71 genes had differential expression in BEL-7402 cell line. 24 genes were up-regulated while 7 genes were down-regulated. In those genes, many differentially expressed genes were involved in cell division, membrane receptor and immunocompeten. Abnormal gene expression contribute to malignant cell divison. Cell survive rates and activities were detected by typan blue exclusion respectively, phase contrast microscopy , flowcytometer and cDNA microarray were used to detect the ability to inhibit the growth of BEL-7402 in vitro. It may be an important mechanism for Somatostatin inhibition of proliferative ability of BEL-7402 cells by inducing cell differentiation and modulating the cell cycle progress, rather than by killing the hepatocellular carcinoma cell or inducing apoptosis directly. Nude mice bearing xenografts of cancer cell line were treated with Somatostatin, Somatostatin and Interferon-alpha or saline control for 4 weeks after tumor implantation .The mechanism of Inhibition of Somatostatin on proliferative ability of tumor cells was observed bymeasuring tumor volume, MVD were detected with immunohistochemistry in three groups. Cell cycle progress and apoptosis were detected by flowcytometer, gene expression profile was identified by cDNA microarrays. The ratio of cell in resting state (G0/G1) increased, MVD decreasing , apopotosis peak was observed, there were 23 genes had differential expression. 7 genes were up-regulated while 16 genes were down-regulated, specific genes involved in modulating the cell cycle progress , apoptosis cascade. These finding suggest that the inhibition on effects occur in the growth of HCC transplant tumor in nude mice were markedly improved by Somatostatin combined with Interferon-alpha when compared with Somatostatin alone. To administrate Somatostatin was security after resection of liever, it could enhance wound healing. These synergic results may be of potential therapeutic benefit to those patients with HCC.更多还原