SIGNAL TRANSDUCTION IN NORMAL AND TRANSFORMED OSE

卵巢癌是恶性肿瘤中最具侵袭性和转移性的形式之一。 女性生殖器癌症，导致死亡率过高 与发病率相比。尽管流行病学研究表明 与排卵和反复伤口愈合相关的增殖性事件， 关于调节增殖的过程知之甚少， 正常人卵巢表面上皮（OSB）或恶性卵巢上皮细胞 上皮细胞研究正常人OSE的主要障碍之一是 获得足够的细胞数量， 分子和生物化学研究。我们使用了一种新开发的生产线 SV 40 T抗原转染的人OSE细胞的增殖能力增加， 研究甚至与基因表达和DNA相关的信号传导的潜力 在OSB中合成。这些细胞的表型正常， 通过在裸鼠中的锚定依赖性生长和缺乏致瘤性。我们 初步结果表明，蛋白激酶C（pKC）的激活 导致OSE中DNA合成的快速和持续抑制 细胞，这种抑制作用进一步增加毒胡萝卜素处理 以及随后的细胞内钙的释放。观察到的 正常OSB细胞中DNA合成的抑制被一种 抑制细胞增殖。相反，SKOV-3中的DNA合成 卵巢癌细胞系对PKC的短期效应不太敏感 活化和用佛波醇酯长期处理SKOV-3细胞 肿瘤促进剂12-0-十四酰基-13-佛波醇乙酸酯CrpA）实际上 导致DNA合成增加，可能是由于 活性PKC这些观察结果使我们假设PKC是一种 OSB细胞增殖的重要调节因子， PKC相关的信号转导通路可能与 转化细胞中生长失控。我们还注意到， OSE细胞中的PKC激活导致快速和持续的 JunB mRNA表达以及c-Jun和c-Fos的瞬时诱导 mRNA。我们将检验PKC诱导JunB蛋白变化的假设， 表达介导的PKC激活的影响，通过观察 JunB缺失和过表达对OSB细胞DNA合成的影响。我们 还将确定PKC激活对磷酸化的影响， Jun和Fos蛋白质。我们将描述PKC之间的相互作用 和细胞内钙，以及PKC亚型的表达在正常人 和转化的OSE细胞。如果这些研究结果支持我们的观点， PKC相关信号转导的实质性差异假说 正常细胞和恶性细胞之间的通路，我们将测试假设 过度表达适当的PKC亚型， 细胞，观察其对裸鼠成瘤性的影响。这些 研究应提供有关两个主要信号的重要信息 转导系统，可能有助于调节 OSE细胞的增殖和分化。

Ovarian carcinoma is one of the most aggressive and metastatic forms of female genital cancer, resulting in a disproportionately high death rate compared to incidence. Although epidemiological studies have implicated proliferative events associated with ovulation and repeated wound healing, very little is known about the processes regulating proliferation in either normal human ovarian surface epithelium (OSB) or malignant ovarian epithelial cells. One of the major barriers to studies of normal human OSE cells has been the difficulty in obtaining sufficient cell numbers for molecular and biochemical studies. We have used a recently developed line of SV4O T-antigen transfected human OSE cells with increased proliferative potential to study signaling even associated with gene expression and DNA synthesis in the OSB. These cells are phenotypically normal as evidenced by anchorage dependent growth and lack of tumorigenicity in nude mice. Our preliminary results indicate that activation of protein kinase C (pKC) results in a rapid and persistent inhibition of DNA synthesis in OSE cells, and this inhibition is further increased by thapsigargin treatment and the subsequent release of intra cellular calcium. The observed inhibition of DNA synthesis in normal OSB cells is paralleled by an inhibition of cell proliferation. In contrast, DNA synthesis in the SKOV-3 ovarian tumor cell line is less sensitive to the short term effects of PKC activation, and chronic treatment of SKOV-3 cells with the phorbol ester tumor promoter 12-0-tetradecanoyl-13-phorbol acetate CrpA) actually results in an increase in DNA synthesis, probably due to the depletion of active PKC. These observations have led us to hypothesize that PKC is an important modulator of proliferation in OSB cells, and that changes in PKC-associated signal transduction pathways may be causally related to the loss of growth control in transformed cells. We have also observed that PKC activation in OSE cells results in a rapid and sustained induction of JunB mRNA expression, as well as transient inductions of c-Jun and c-Fos mRNA. We will test the hypothesis that PKC-induced changes in JunB protein expression mediate the effects of PKC activation by observing the effects of JunB depletion and overexpression on DNA synthesis in OSB cells. We will also determine the effects of PKC activation on the phosphorylation of Jun and Fos proteins. We will characterize the interactions between PKC and intracellular calcium, and the expression of PKC isoforms in normal and transformed OSE cells. If the results of these studies support our hypothesis of substantial differences in the PKC-associated signaling pathways between normal and malignant cells, we will test the hypothesis of causality by overexpressing the appropriate PKC isoform in transformed cells and observing the effects on tumorigenicity in nude mice. These studies should provide important information about two major signal transduction systems which may contribute to the regulation of proliferation and differentiation in OSE cells.