口腔鳞癌癌细胞和癌相关成纤维细胞的原代培养、鉴定及其生物学特性研究

目的：对口腔鳞状细胞癌细胞（OSCCs）及癌相关成纤维细胞（CAFs）进行培养、纯化和鉴定,初步探讨其生物学特性。方法：取自临床手术的新鲜标本,通过抗污染处理,应用机械分离法和胶原酶Ⅳ消化法获得OSCC及CAFs,0.25%胰蛋白酶消化反复贴壁差数培养法分离、纯化细胞,正常口腔成纤维细胞（NFs）和舌鳞癌细胞系Tca8113作为对照组。通过形态学观察、波形蛋白和角蛋白免疫组化染色对纯化的细胞进行鉴定。应用MTT法绘出OSCCs、CAFs、NFs和Tca8113细胞的生长曲线。结果：获得纯化的OSCCs和CAFs,通过对比研究阐明OS-CC和CAFs的生物学特点;OSCCs中免疫组化染色角蛋白呈阳性、波形蛋白呈阴性,CAFs中角蛋白呈阴性、波形蛋白呈阳性。结论：通过本实验研究成功获得纯化的OSCCs和CAFs,对比研究发现CAFs和NFs在形态结构、生长特点等方面不同。

Primary Culture and Identification of Oral Squamous Carcinoma Cells and Carcinoma-associated Fibroblasts in vitro, and Their Bionomics

Objective：To cultivate,purify and identify oral squamous carcinoma cells（OSCCs）and carcinoma-associated fibroblasts（CAFs）,and investigate their bionomics preliminarily.Methods：The primary oral squamous carcinoma tissues were obtained from fresh specimen tissue.Then cells were dissociated by mechanical separating method and collagenase typeⅣdigestiving method.Cells were digestived by 0.25% trypsin and were purified by adherence again and again.Morphological characteristics were observed under light microscope.Oral normal fibrolasts and oral squamous carcinoma cells line（Tca 8113）were set as the control group.The proteins including vimentin and cytokeratin were examined by immunohistochemistry.Proliferation was tested with method.Results：The purified oral CAFs and OSCCs were maintained.The biological characteristics of OSCC and CAFs were expounded by comparing to the control.The oral CAFs showed negative staining for cytokeratin,and positive staining for vimentin,and the OSCCs showed positive staining for cytokeratin,and negative staining for vimentin.Conclusion：The purified OSCCs and CAFs were obtained in these empirical study.There were significant differences on the morphological characteristics and growth feature between the CAFs and NFs.