【作者】 欧阳运薇；

【导师】 彭芝兰；

【作者基本信息】 四川大学 ， 妇产科学， 2005， 博士

【摘要】 目的:本研究拟通过检测肿瘤转移抑制基因KAI1/CD82在宫颈癌组织中的蛋白表达状况,以及观察它对宫颈癌Caski细胞的生物学特性的影响,对该基因与宫颈癌侵袭和转移的相关性作一初步探讨,通过检测人乳头瘤病毒HPV16E6、E7,HPV18在宫颈癌组织中的感染情况,探讨HPV是否影响KAI1/CD82基因的表达。 方法:采用免疫组化LsAB法对70例浸润性宫颈癌,15例原位癌,20例正常宫颈组织以及29例转移淋巴结的石蜡标本进行KAI1/CD82蛋白表达检测,分析KAI1/CD82与宫颈癌临床分期、病理分级、病理类型、淋巴结转移、宫颈间质肌层浸润深度等病理特点的关系;采用PCR扩增-琼脂糖凝胶电泳法检测浸润性宫颈癌以及原位癌组织中人乳头瘤病毒HPV16E6、E7,HPV18的DNA状况;分析HPV16E6、E7,HPV18感染与KAI1/CD82蛋白表达的相关性;通过脂质体转染技术,将真核表达质粒PCMV-KAI1 cDNA导入低表达的宫颈癌高转移性细胞系Caski,通过MTT法测生长曲线以及粘附实验、侵袭力小室观察细胞体外生长、体外侵袭能力以及对人工细胞外基质的粘附情况。 结果:1.KAI1/CD82在宫颈浸润癌组织及原位癌中的蛋白表达与正常宫颈上皮中的表达相比明显下调(P<0.05),主要位于上皮细胞的胞膜,在癌细胞中还表达于胞浆,在癌组织中以不均匀性灶性分布为主,更多还原

【Abstract】 Objective:To research the expression of KAI1/CD82 metastasis suppressor gene in cervical carcinoma and study the effect of kail gene on the proliferation properties and invasive ability of Caski cervical cancer cell transfected by recombinant plasmid pCMV-KAI1 in vitro;also study the infection of HPV16E6, E7and HPV18 in cervical carcinoma to explore its effect on the expression of kail.Methods:Immunohistochemistry was used to detect the expression of KAI1/CD82 gene in formalin-fixed,paraffin-embedded specimens of 20 normal cervical epthelium,15 cervical in situ carcinoma,70 primary invasive cervical carcinoma as well as 29 lymph node metastases,Correlation of expression of KAI1 gene with clinicopathologic factors were statistically analyzed.The status of HPV16 E6, E7and HPV18 DNA was detected by PCR method; PCMV-KAI1 cDNA was transfected into highly malignant cervical carcinoma cell line CaSki by liposome,which had low levels of endogenous KAI1 expression .The expression of KAI1 protein was determined by immunohistochemistry,and the biological behaviors of the KAI1-transfected Caski cells were investigated by cell-matrix adhension and invasion assays.ResultS:l.KAIl/CD82 was expressed in both tumor and normal cervical epthelium, chiefly located in cell membrane,also locatecd in plasma in cancer cells; KAIl protein expression in tumor cells were down-regulation in invasive carcinoma and in situ carcinoma compared to controls(P<0.05).2.The protein expression of KAIl was not correlated with tumor FIGO stage> pathological subt)^^ depth of cervical infiltration^ lymphovascular space invasion .and the pelvic lymph node metastasis of cervical carcinoma(P>0.05),but its expression was significantly decreased in poorly differentiated tumours compared to that of well and moderately differentiated tumours,KAIl also expressed in lymph node metastases,but there have no statistically significant difference between them with expression in situ.3.The infection of HPV16 E6, E7and HPV18 in invasive carcinoma was not higher than in situ carcinoma , also not correlated with tumor clinical stage,histological grade^ depth of cervical infiltration lymphovascular space invasion and the pelvic lymph node metastasis,The expression of HPV16E6 was correlated with squamous cervical carcinoma,there was no correlation between expression of KAIl with infection of HPV16 E6, E7 or HPV18.4.The level of protein expression of Kail gene in Caski-kail cell after transfected by recombinant plasmid pCMV-KAIl was strengthed proved by Immunohistochemistry and RT-PCR(P<0.05).5. After transfected by recombinant plasmid pCMV-KAIl in vitro ,The proliferation ability of Caski cervical cancer cell were suppresssed compared with Caski and Caski-vector cell transfected by vector, and the migrative ability of passing through the membrane filte also decreased evidently(P<0.05).6.The ability of adhension of Caski-Kail cell to extracellular matrix was enhanced after trasfected with KAIl gene compared with Caski and Caski-vector cell,but there have no difference between Caski cell andCaski-vector cell (P>0.05) .Conclusion: The KAI1/CD82 metastasis suppressor gene might not a main factor in invasion and metastasis of cervical carcinoma , rough it affect the ability of proliferation and invasion of cervical cancer cell Caski in vitro.HPV infection doesn’t affect the expression of KAI1 gene.更多还原