Pietras R J, Szego C M
J Cell Biol. 1979 Jun;81(3):649-63. doi: 10.1083/jcb.81.3.649.
Endometrial cells isolated from uteri of ovariectomized rats were treated in vitro with 1 X 10(-9) M estradiol-17 beta (E2beta) to analyze early changes in membrane properties during hormone-induced growth. After 30-min exposure to E2beta at 22 degrees C, cells exhibited an enhanced capacity to bind erythrocytes (hemadsorption) in the presence of concanavalin A (Con A) to 237% of the level in paired controls. Fluorescence microscopy revealted that approximately 25% of cells exposed to E2beta, but not estradiol-17 alpha (E2alpha), showed a redistribution into polar clusters of Con A-binding sites that were dispersed in random patches at the external surfaces of control cells. These hormore-induced membrane alterations were abolished by prior treatment of cells with inhibitors of thiol proteinase activity of the cathepsin B1 (CB1) type, such as leupeptin and iodoacetate. Leupeptin at 4.5 X 10(-7) M also reduced the affinity of [3H]E2beta binding to intact cells but did not influence specific binding of the hormone to macromolecular components of cytosol. A pronounced increase in the availability of endogenous CB1, But not of alkaline phosphatase, succinate, or lactate dehydrogenase, in the extracellular media was elicited within 30 min after E2beta treatment. In cells cultured in chemically defined medium for up to 48 h, E2beta, but not E2alpha, enhanced cell proliferation and stimulated [3H]thymidine incorporation into macromolecular form. These E2beta-induced effects were abolished by prior treatment of cells with liposome-entrapped leupeptin at a final concentration of 7 X 10(-8) M. The net rate of intercellular adhesion among endometrial cells was also enhanced by E2beta. This hormonal response was diminished by prior exposure to leupeptin. Fractionation of cells by selection for adhesiveness due to E2beta exposure for 30 min yielded a subpopulation of rapidly dividing cells which surpassed their less adhesive counterparts in cathepsin secretion and in Con A-mediated hemadsorption. These results indicate that leupeptin-sensitive proteinase activity may contribute to membrane and growth modifications elicited by E2beta treatment in endometrial cells.
从去卵巢大鼠子宫中分离出的子宫内膜细胞，在体外以1×10⁻⁹ M的17β-雌二醇（E2β）处理，以分析激素诱导生长过程中膜特性的早期变化。在22℃下暴露于E2β 30分钟后，细胞在伴刀豆球蛋白A（Con A）存在下结合红细胞（血细胞吸附）的能力增强，达到配对对照组水平的237%。荧光显微镜显示，约25%暴露于E2β而非17α-雌二醇（E2α）的细胞，其Con A结合位点重新分布成极性簇，而在对照细胞外表面这些位点呈随机斑块状分布。这些激素诱导的膜改变可通过用组织蛋白酶B1（CB1）型巯基蛋白酶活性抑制剂（如亮抑酶肽和碘乙酸盐）预先处理细胞而消除。4.5×10⁻⁷ M的亮抑酶肽也降低了[³H]E2β与完整细胞结合的亲和力，但不影响该激素与胞质溶胶大分子成分的特异性结合。E2β处理30分钟后，细胞外培养基中内源性CB1的活性显著增加，而碱性磷酸酶、琥珀酸或乳酸脱氢酶的活性未增加。在化学成分确定的培养基中培养长达48小时的细胞中，E2β而非E2α增强了细胞增殖并刺激[³H]胸苷掺入大分子形式。这些E2β诱导的效应可通过用终浓度为7×10⁻⁸ M的脂质体包裹的亮抑酶肽预先处理细胞而消除。E2β还增强了子宫内膜细胞间的净黏附率。预先暴露于亮抑酶肽可减弱这种激素反应。通过选择因暴露于E2β 30分钟而具有黏附性的细胞进行分级分离，得到了一个快速分裂的亚群细胞，其在组织蛋白酶分泌和Con A介导的血细胞吸附方面超过了黏附性较低的对应细胞。这些结果表明，亮抑酶肽敏感的蛋白酶活性可能有助于E2β处理诱导的子宫内膜细胞膜和生长变化。
