Stein L S, Stoica G, Tilley R, Burghardt R C
Department of Veterinary Anatomy, Texas A&M University, College Station 77843.
Cancer Res. 1991 Jan 15;51(2):696-706.
A spontaneously immortalized clonal granulosa cell line (SIGC) derived from primary rat ovarian granulosa cell cultures was developed as a model system to explore the process of transformation using an epithelial cell type. SIGC has an epithelial morphology and grows in culture without undergoing luteinization. The cell line is thought to represent an intermediate step in carcinogenesis because it seems to grow indefinitely in culture but does not form clones in soft agar or tumors in nude mice. Indirect immunofluorescence and Western blot analysis verified the constitutive expression of the recessive oncogene product p53 in the cell line, thereby suggesting a possible mechanism of immortalization. Ultrastructural studies indicated that SIGC cells are characterized by an undifferentiated phenotype with prominent intermediate filaments, desmosomes, and gap junctions. The identification of cytokeratin by indirect immunofluorescence and Western blot analysis suggests that SIGC functions as an epithelial cell type. Functional studies of cell-cell communication by a dye transfer technique (fluorescence recovery after photobleaching) showed reduced communication compared to normal primary granulosa cells in culture. SIGC cells were transfected with early region genes of SV40 virus in an attempt to generate fully transformed cell lines. The resulting cell line SV-SIGC expressed T-antigen, was anchorage independent, formed tumors in nude mice, and had reduced intercellular communication as compared to SIGC cells. Explants from the tumors in nude mice were used to generate another cell line (T-SV-SIGC), which exhibited further reduction in both the incidence and the rate of communication. These results clearly demonstrated a progressive loss of functional communication during multistep transformation of an ovarian cell type. These data demonstrate that this assay system based on an epithelioid cell type can be used to study the relationship between intercellular communication and the multistep process of carcinogenesis.
从原代大鼠卵巢颗粒细胞培养物中获得了一种自发永生化的克隆颗粒细胞系（SIGC），将其作为一个模型系统，以利用上皮细胞类型来探索细胞转化过程。SIGC具有上皮形态，在培养中生长且不发生黄体化。该细胞系被认为代表了癌变过程中的一个中间步骤，因为它似乎能在培养中无限生长，但在软琼脂中不形成克隆，在裸鼠中也不形成肿瘤。间接免疫荧光和蛋白质印迹分析证实了该细胞系中隐性癌基因产物p53的组成型表达，从而提示了一种可能的永生化机制。超微结构研究表明，SIGC细胞的特征是具有未分化的表型，有明显的中间丝、桥粒和缝隙连接。通过间接免疫荧光和蛋白质印迹分析鉴定细胞角蛋白，表明SIGC作为一种上皮细胞类型发挥功能。通过染料转移技术（光漂白后荧光恢复）对细胞间通讯进行的功能研究表明，与培养中的正常原代颗粒细胞相比，通讯减少。用SV40病毒的早期区域基因转染SIGC细胞，试图产生完全转化的细胞系。所得的细胞系SV - SIGC表达T抗原，不依赖贴壁生长，在裸鼠中形成肿瘤，与SIGC细胞相比，细胞间通讯减少。用裸鼠肿瘤的外植体产生了另一个细胞系（T - SV - SIGC），其通讯发生率和通讯速率进一步降低。这些结果清楚地证明了在卵巢细胞类型的多步骤转化过程中功能通讯的逐渐丧失。这些数据表明，这种基于上皮样细胞类型的检测系统可用于研究细胞间通讯与癌变多步骤过程之间的关系。
