Jiang Feng, Saunders Beatriz O, Haller Edward, Livingston Sandra, Nicosia Santo V, Bai Wenlong
Department of Pathology, University of South Florida College of Medicine and H. Lee Moffitt Cancer Center, Tampa, Florida 33612-4799, USA.
In Vitro Cell Dev Biol Anim. 2003 Jul-Aug;39(7):304-12. doi: 10.1290/1543-706X(2003)039<0304:CIOCLF>2.0.CO;2.
The tendency of the ovarian surface epithelium (OSE) to undergo metaplastic and morphogenetic changes during the life cycle, at variance with the adjacent peritoneal mesothelial cells, suggests that its biology may be regulated by underlying ovarian stromal cues. However, little is known about the role that the ovarian stroma plays in the pathobiology of the OSE, largely because of the lack of a suitable in vitro model. Here, we describe the establishment and characterization of conditionally immortalized ovarian stromal and surface epithelial cell lines from H-2K(b)-tsA58 transgenic mice that carry the thermolabile mutant of SV-40 large T antigen under the control of an interferon-gamma (IFN-gamma)-inducible promoter. These cells express functional T antigens, grow continuously under permissive conditions at 33 degrees C in the presence of IFN-gamma, and stop dividing when the activity and expression of the tumor antigen is suppressed by restrictive conditions without IFN-gamma at 39 degrees C. Morphological, immunohistochemical, and ultrastructural analyses show that conditionally immortal OSE cells form cobblestone-like monolayers, express cytokeratin and vimentin, contain several microvilli, and develop tight junctions, whereas stromal cells are spindle-like, express vimentin but not cytokeratin, and contain rare microvilli, thus exhibiting epithelial and stromal phenotypes, respectively. At variance with the reported behavior of rat epithelial cells, conditionally immortal mouse epithelial cells are not spontaneously transformed after continuous culture in vitro. More importantly, conditioned media from stromal cells cultured under permissive conditions increase the specific activity of the endogenous estrogen receptor in BG-1 human ovarian epithelial cancer cells and promote these cells' anchorage-independent growth, suggesting the paracrine influence of a stromal factor. In addition, stromal cells cultured under restrictive conditions retain this growth-stimulatory activity, which, therefore, appears to be independent of T antigen expression. These established cell lines should provide a useful in vitro model system for studying the role of cellular interactions in OSE cell growth and tumorigenesis.
卵巢表面上皮（OSE）在生命周期中经历化生和形态发生变化的倾向，与相邻的腹膜间皮细胞不同，这表明其生物学特性可能受潜在的卵巢基质信号调节。然而，关于卵巢基质在OSE病理生物学中所起的作用知之甚少，这主要是因为缺乏合适的体外模型。在此，我们描述了从H-2K(b)-tsA58转基因小鼠建立并鉴定条件性永生化卵巢基质和表面上皮细胞系，这些小鼠在干扰素-γ（IFN-γ）诱导型启动子的控制下携带SV-40大T抗原的温度敏感突变体。这些细胞表达功能性T抗原，在33℃允许条件下，在IFN-γ存在时持续生长，而当肿瘤抗原的活性和表达在39℃无IFN-γ的限制条件下被抑制时停止分裂。形态学、免疫组织化学和超微结构分析表明，条件性永生化OSE细胞形成鹅卵石样单层，表达细胞角蛋白和波形蛋白，含有多个微绒毛，并形成紧密连接，而基质细胞呈纺锤形，表达波形蛋白但不表达细胞角蛋白，含有稀少微绒毛，因此分别表现出上皮和基质表型。与报道的大鼠上皮细胞行为不同，条件性永生化小鼠上皮细胞在体外连续培养后不会自发转化。更重要的是，在允许条件下培养的基质细胞的条件培养基增加了BG-1人卵巢上皮癌细胞中内源性雌激素受体的比活性，并促进这些细胞的非贴壁依赖性生长，提示基质因子的旁分泌影响。此外，在限制条件下培养的基质细胞保留了这种生长刺激活性，因此，这种活性似乎与T抗原表达无关。这些建立的细胞系应为研究细胞间相互作用在OSE细胞生长和肿瘤发生中的作用提供一个有用的体外模型系统。
