Tanahashi Kayo, Shibahara Shinobu, Ogawa Minako, Hanazono Makoto, Aizawa Shinichi, Tomooka Yasuhiro
Department of Biological Science and Technology, Tokyo University of Science, 2641 Yamazaki Noda, Chiba 278-8510, Japan.
In Vitro Cell Dev Biol Anim. 2002 Nov-Dec;38(10):547-56. doi: 10.1290/1543-706x(2002)38<547:eacocc>2.0.co;2.
Clonal cell lines have been established from vagina of prepubertal female p53(-/-) mice. Because the mouse vagina has a dual origin (the cranial three-fifths derived from the Müllerian duct and the caudal two-fifths derived from the urogenital sinus), both parts were separately subjected to cloning. Sixteen epithelial and two fibroblastic cell lines were established from the cranial three-fifths (Müllerian vagina group), and four epithelial and three fibroblastic cell lines were established from the caudal two-fifths (sinus vagina group). They were maintained in Dulbecco's modified Eagle medium and Ham's nutrient mixture F-12 containing 10% fetal calf serum and 17 beta-estradiol at 10(-8) M. Two cell lines (one epithelial and one fibroblastic) were examined using soft agar assay, but no colonies were formed. The doubling time of the cell lines was approximately 24 h, and all of them divided more than 200 times without crisis, suggesting that they were immortalized. All epithelial cell lines expressed cytokeratin 8. However, the epithelial cell lines expressed cytokeratin 14 and cytokeratin 10 when exposed to medium containing different concentrations of Ca(2+). Fibroblastic cell lines expressed vimentin. All epithelial and fibroblastic cell lines expressed estrogen receptor-alpha protein. This is the first successful establishment of clonal cell lines from the normal mouse vagina, and these lines may provide good models in vitro of the vagina for the study of the mechanism of estrogen action.
已从青春期前雌性 p53(-/-) 小鼠的阴道中建立了克隆细胞系。由于小鼠阴道有双重起源（前三分之五源自苗勒管，后五分之二源自泌尿生殖窦），这两部分分别进行了克隆。从阴道的前三分之五（苗勒阴道组）建立了 16 个上皮细胞系和 2 个成纤维细胞系，从后五分之二（窦阴道组）建立了 4 个上皮细胞系和 3 个成纤维细胞系。它们在含有 10%胎牛血清和 10(-8) M 的 17β-雌二醇的 Dulbecco 改良 Eagle 培养基和 Ham 营养混合物 F-12 中培养。使用软琼脂试验检测了两个细胞系（一个上皮细胞系和一个成纤维细胞系），但未形成集落。这些细胞系的倍增时间约为 24 小时，并且所有细胞系均分裂超过 200 次而无危机，表明它们已永生化。所有上皮细胞系均表达细胞角蛋白 8。然而，当暴露于含有不同浓度 Ca(2+) 的培养基中时，上皮细胞系表达细胞角蛋白 14 和细胞角蛋白 10。成纤维细胞系表达波形蛋白。所有上皮和成纤维细胞系均表达雌激素受体-α 蛋白。这是首次成功从正常小鼠阴道中建立克隆细胞系，这些细胞系可能为体外研究雌激素作用机制提供良好的阴道模型。
