Proliferation of mouse uterine epithelial cells in vitro.
Proliferation of mouse uterine epithelial cells in vitro.
复制标题
小鼠子宫上皮细胞的体外增殖。
作者：
Y. Tomooka;R. Diaugustine;J. Mclachlan
虽然雌激素可以刺激子宫上皮细胞在体内的生长，但雌激素对生殖道组织上皮细胞的体外生长没有明确的影响;因此，我们建立了一个明确的小鼠子宫上皮细胞培养系统。用胰蛋白酶处理未成熟CD-1小鼠（21-23日龄）的子宫碎片，分离上皮碎片，通过Percoll梯度离心富集，并接种在从大鼠尾腱制备的胶原凝胶上。最初，将细胞培养在Ham’s F-12和Dulbecco’s Modified Eagle’s培养基的1：1混合物中，该混合物补充有表皮生长因子（EGF; 10 ng/ml）、胰岛素（10微克/ml）、转铁蛋白（10微克/ml）、氢化可的松（0.1微克/ml）和维生素A（10 ng/ml）。细胞在1-2天内在胶原凝胶上形成单层，但随着时间的推移，细胞开始从凝胶上脱离。进一步的研究表明，这些细胞在胶原上的附着和生长受到钙浓度的显著影响。发现将钙浓度从1.05降低至0.05-0.1 mM显著抑制细胞脱离;培养7天后细胞数量加倍。EGF可促进子宫上皮细胞的增殖，而成纤维细胞生长因子、血小板衍生生长因子、神经生长因子、增殖刺激活性或生长调节素C则不能。子宫上皮细胞对[125 I]碘-EGF（Kd，约1.8 nM）显示出单一类别的高亲和力结合位点，约5 × 10（4）个受体/细胞;结合被EGF抑制，但不被其他多肽抑制。这种细胞培养系统将有助于我们研究激素对雌激素靶细胞生长和分化的影响。
Although estrogens can stimulate the growth of uterine epithelial cells in vivo, there is no clear effect of estrogens on the in vitro growth of epithelial cells from reproductive tract tissues; thus, we have established a defined culture system for mouse uterine epithelial cells. Pieces of uteri from immature CD-1 mice (21-23 days of age) were treated with trypsin, and the epithelial fragments were separated, enriched by Percoll gradient centrifugation, and seeded on collagen gels prepared from rat tail tendon. Initially, the cells were cultured in a 1:1 mixture of Ham's F-12 and Dulbecco's Modified Eagle's Medium supplemented with epidermal growth factor (EGF; 10 ng/ml), insulin (10 micrograms/ml), transferrin (10 micrograms/ml), hydrocortisone (0.1 micrograms/ml), and vitamin A (10 ng/ml). The cells formed a monolayer on the collagen gel within 1-2 days, but with time, cells began to detach from the gel. Further studies revealed that the attachment and growth of these cells on collagen were markedly influenced by the calcium concentration. It was found that lowering the calcium concentration from 1.05 to 0.05-0.1 mM dramatically suppressed cell detachment; the number of cells doubled after 7 days of culture. Proliferation of uterine epithelial cells was enhanced by EGF, but not by fibroblast growth factor, platelet-derived growth factor, nerve growth factor, multiplication-stimulating activity, or somatomedin-C. The uterine epithelial cells exhibited a single class of high affinity binding sites for [125I]iodo-EGF (Kd, approximately 1.8 nM), with approximately 5 X 10(4) receptors/cell; binding was inhibited by EGF but not by the other polypeptides. This cell culture system will aid in our investigations on hormonal effects on the growth and differentiation of estrogen target cells.