Molecular biologic study on the growth inhibition of human endometrial glands.

人类子宫内膜腺细胞的生长被认为是由雌激素刺激的，而被黄体酮抑制的。在本研究中，为了从细胞周期调节机制的角度了解黄体酮对人子宫内膜腺细胞生长的抑制作用，我们进行了以下实验。(1)细胞周期蛋白和细胞周期蛋白依赖激酶（cyclin-dependent kinase, cdks）的免疫组化表达在增殖期，而在分泌期未见表达。在几种肿瘤抑制基因产物中，只有p27在分泌期被免疫组织化学观察到。(2)免疫印迹法检测体外培养的子宫内膜腺细胞中cyclins和cdks的表达在黄体酮处理后降低。相反，在黄体酮处理的细胞中，p27蛋白的表达呈剂量依赖性增加。(3) Northern blotting检测p27mRNA的表达在增殖期和分泌期之间无显著差异。(4)为评价黄体酮对体外培养子宫内膜腺p27蛋白水解的影响，在诱导环己亚胺（一种蛋白质合成抑制剂）后每12小时（最长48小时）用免疫印迹法检测黄体酮处理和不处理黄体酮的子宫内膜腺p27蛋白的表达量。结果表明，在孕激素处理的细胞中，p27蛋白的含量在每个时间过程中都更丰富。(5) p27表达质粒在体外培养的子宫内膜腺中强制表达p27蛋白，诱导p27表达增加，增殖细胞核抗原（增殖指标PCNA）表达降低。综上所述，本研究表明黄体酮诱导的细胞周期蛋白/cdks的表达降低以及p27的表达升高可能参与了子宫内膜腺的生长抑制。

The growth of the glandular cells of the human endometrium is thought to be stimulated by estrogen, and inhibited by progestin. In the present study, to understand the progesterone-induced growth inhibition of the glandular cells of the human endometrium in a view of cell cycle regulatory machineries, following experiments ,were performed. (1) The immunohistochemical expression of cyclins and cyclin-dependent kinases (cdks) of the glandular cells was observed in the proliferative phase, but not in the secretory phase. Among several tumor suppressor gene products, only p27 was immunohistochemically observed in the secretory phase. (2) The expression of cyclins and cdks detected by immunoblotting in cultured endometrial glandular cells was reduced in progesterone treated cells. Conversely, the expression of p27 protein was increased in a dose dependent fashion in those cells treated with progesterone. (3) The expression of p27mRNA examined by Northern blotting analysis did not vary significantly between the proliferative and the secretory phases. (4) To evaluate the effect of progesterone on the proteolysis of p27 in cultured endometrial glands, amount of p27 protein treated with and without progcsterone was evaluated by immunoblotting in every 12 hours (up to 48 hours) after cycloheximide (a protein synthesis inhibitor) was induced. The result showed that the amount of p27 protein was more abundant in those cells treated with progesterone in every time course examined. (5) Forced expression of p27 protein by p27 expression plasmid in cultured endometrial glands induced increased expression of p27 and reduced expression of proliferating cell nuclear antigen (PCNA, an indicator of cell proliferation). In summary, the present study demonstrated that progesterone-induced reduced expression of cyclins/cdks, as well as elevated expression of p27 may be involved in the growth inhibition of endometrial glands.