SIGNAL TRANSDUCTION IN NORMAL AND TRANSFORMED OSE

卵巢癌是最具侵袭性和转移性的卵巢癌之一。 女性生殖器癌，导致不成比例的高死亡率 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， 人们对核扩散的调控过程知之甚少。 正常卵巢表面上皮(OSB)或恶性卵巢 上皮细胞。阻碍正常人OSE研究的主要障碍之一 细胞一直是获得足够的细胞数量的困难 分子和生化研究。我们用的是最近开发的一条线路 SV4O T抗原转染人OSE细胞增殖增强的研究 研究甚至与基因表达和DNA相关的信号的可能性 在OSB中进行综合。这些细胞在表型上是正常的 在裸鼠体内依靠锚定生长和缺乏致瘤性。我们的 初步结果表明，蛋白激酶C(PKC)的激活 导致OSE中DNA合成的快速和持久的抑制 细胞，这种抑制作用被thapsigargin处理进一步增强 以及随后细胞内钙的释放。被观察到的 An对正常OSB细胞DNA合成的抑制作用 抑制细胞增殖。相比之下，SKOV-3中的DNA合成 卵巢肿瘤细胞系对PKC的短期效应不太敏感 佛波酯对SKOV-3细胞的激活和慢性作用 肿瘤促进剂12-0-十四酰-13-佛波醇醋酸酯(CRPA) 导致DNA合成增加，可能是由于 激活的PKC。这些观察结果使我们假设PKC是一种 OSB细胞中重要的增殖调节因子，而这种变化 PKC相关的信号转导通路可能与 在转化细胞中失去生长控制。我们还观察到， OSE细胞中PKC的激活导致快速而持续的诱导 JunB基因的表达及其对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 表达通过观察PKC的激活效应来介导PKC的激活效应 OSB细胞中JunB缺失和过表达对DNA合成的影响我们 也将决定PKC激活对磷酸化的影响 Jun和Fos蛋白。我们将描述PKC之间的相互作用 和细胞内钙，以及PKC亚型在正常组织中的表达 和转化的OSE细胞。如果这些研究的结果支持我们的 PKC相关信号存在显著差异的假说 正常和恶性细胞之间的通路，我们将检验这一假说 通过过度表达适当的PKC亚型来研究因果关系 细胞，并观察其对裸鼠致瘤性的影响。这些 研究应该提供关于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.