经绿色荧光蛋白基因修饰的人脉络膜黑色素瘤细胞株OCM-1-gfp的生物学行为与基因表达

目的 研究经绿色荧光蛋白（GFP）基因修饰的人脉络膜黑色素瘤细胞株OCM-1-gfp在小鼠体内的生长过程、转移规律，以及可能影响肿瘤生长和转移的因素。 方法 用脂质体将GFP基因导入人脉络膜黑色素瘤细胞OCM-1，建立稳定、高水平表达GFP的克隆;分别接种到Balb/c裸鼠视网膜下和后大腿皮下，建立原位和异位肿瘤模型。眼内肿瘤生长情况用荧光显微镜直接观察，皮下肿瘤大小用游标卡尺测量;采用免疫组织化学方法对肿瘤内13种基因的表达进行检测。 结果 稳定表达GFP的脉络膜黑色素瘤细胞OCM-1-gfp基本保持了亲代细胞的特征;能在裸鼠体内形成肿瘤并继续生长和转移;在基因表达方面，肿瘤抑制基因p16染色呈阴性，p53染色呈强阳性。其他的肿瘤抑制基因:视网膜母细胞瘤易感基因（Rb）、p21，转录调控因子（E-2F）、核因子κB（NFκB），细胞增生相关基因细胞周期素D1（cyclin D1）、增生细胞核抗原（PCNA），细胞凋亡相关基因bcl-2、bcl-XL/S和bax，以及表皮生长因子（EGF）及其受体（EGFR）呈现不同程度的阳性表达。 结论 GFP为直接观察脉络膜黑色素瘤在体内的生长和转移提供了标记;脉络膜黑色素瘤原位与异位移植瘤模型在成瘤率和生长情况方面无明显差异;由OCM-1-gfp生长形成的肿瘤p16、p53及NFκB、cyclin D1、PCNA及EGF和EGFR等多种基因表达异常。 (中华眼底病杂志, 2006, 22: 170-173)

The biologic behavior and gene expression of malignant choroid melanoma cell strain OCM-1-gfp modified with green fluorescent protein

Objective To investigate the development and metastasis of malignant choroidal melanoma cell strain OCM-1-gfp modified with green fluorescent protein(GFP) and the factors which affected the tumor biological behaviors. Methods GFP was transfected into malignant melanoma cell strain OCM-1.Melanoma cells with high and stable expression of GFP were injected into subretinal space and the subcutaneous space of hind leg of Balb/c nude mouse respectively in order to establish orthotopic and heterotopic transplanted tumor models.The development and metastasis process of orthotopic tumor models was observed directly by fluorescence microscope，and the size of the hypodermal tumor was measured by vernier.The expressions of 13 genes in melanoma were detected by means of immunohistochemistry staining. Results Malignant choroidal melanoma cell strain OCM-1 stably expressed GFP and preserved the characteristics of parental generation，OCM-1-gfp may develop melanoma and continue to metastasize in nude mouse.Positive expression of most of the antibodies，including Rb，p53，p21，E2F，NFκB，cyclin D1，proliferation cellular nuclear antigen（PCNA），bcl 2、bcl XL/S，bax，and epithelial growth factor（EGF）and its receptor（EGFR），was found.While the staining of inhibition gene p16 was negative. Conclusions GFP is the marker for observing the development and metastasis of malignant choroidal melanoma in vivo.The rate of tumor formation and development process in orthotopic models does not differs much from which in heterotopic models of malignant choroidal melanoma.The expressions of lots of genes in malignant choroidal melanoma developed from OCM-1-gfp including p16、p53、NFκB，cyclin D，PCNA，EGF，and EGFR are abnormal. (Chin J Ocul Fundus Dis, 2006, 22: 170-173)