Growth Regulation of the Normal & Malignant Endometrium

正常的生长调节

• 批准号: 6514822
• 负责人: Leslie Ina Gold
• 金额: $ 27.96万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-10 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6514822
• 关键词: adenocarcinoma; athymic mouse; cell cell interaction; cell growth regulation; clinical research; connective tissue cells; cyclin dependent kinase; endometrium; enzyme activity; enzyme inhibitors; epithelium; estrogens; female; gene targeting; genetically modified animals; hormone regulation /control mechanism; hormone related neoplasm /cancer; human subject; mitogen activated protein kinase; mixed tissue /cell culture; progesterone; transcription factor; transforming growth factors; uterus neoplasms; women's health

DESCRIPTION: (provided by applicant) Our broad long-term objectives are to elucidate mechanisms that cause loss of growth inhibition by TGF- b in endometna adenocarcinoma (ECA) and to define hormone regulation of TGF-b through stromal/epithelial interactions in the endometrium. ECA is induced by estrogenic (E2) agents causing hyperproliferation of uterine epithelial cells (UtE). Progesterone (Pg) is therapeutic due its growth inhibitory effect. TGF-b-mediated growth inhibition is transduced by two cooperating receptors (RI, RII) and the downstream signaling/transcription factors, Smad2/3,that activate genes that block cell cycle progression, such as the cyclin-dependent kinase inhibitor, p27kip1 We have shown that UtE isolated from all grades of ECA escape negative growth control by TGF-b by incurring multiple defects in the TGF-b response pathway including, loss of: TGF-b RH, activated Smad2, and p27kip1. Moreover, in complex hyperplasia (CH), the precursor to ECA, these proteins are already decreased. Thus, disruption of TGF-b action occurs early in endometrial carcinogenesis, providing an opportunity to understand molecular events leading to dysregulated growth. We will use primary cultures of normal, CH, and ECA UtE and co-cultures with stromal cells (UtS), which unlike ECA cell lines, retain many in vivo differentiation characteristics. Specific Aim 1, will determine the molecular mechanisms causing TGF-b receptor downregulation (e.g., transcriptional, translational) and test for defective Smad2/3 signaling using a TGF-b-promoter-responsive reporter assay. We will try to regain TGF-b function by transient transfection of RII cDNA into ECA UtE. Specific Aim 2 will test the hypothesis that loss of p27kip1 in ECA is by degradation via ubiquitin-proteasome pathway, which is E2-dnven directly through a MAPkinase via the Ras/MAPK/ERK1 pathway and that TGF-b normally prevents p27 degradation. Tissue/celilysates, inhibitors of proteasomes and MAPK, and immuno-analytical techniques will be used. Using single cell and co-cultures, we show that UtS from normal but not malignant endometnum mediates Pg-induced growth inhibition of normal UtE. Specific Aim 3 will test the hypotheses that UtS paracrine-mediates Pg-induced growth inhibition of UtE by release of TGF-b in response to Pg. We will use co-cultures of normal and "malignant" UtS and UtE and novel in vivo chimeric tissue recombinants composed of UtS from both, Pg receptor knock-out (PRKO) and wild-type mice and human UtE, that are hormonally manipulated as transplants in nude mice and UtE then analyzed for growth. These studies should elucidate molecular mechanisms of endometrial carcinogenesis, hormonal (dys)regulation of endometrial growth, and identify targets for prevention and therapeutic intervention.

描述：(申请人提供)我们的长期目标是 阐明转化生长因子-β对小鼠骨肉瘤生长抑制丧失的机制 子宫内膜腺癌与转化生长因子-β的激素调节 通过子宫内膜中间质/上皮的相互作用。ECA由以下因素引起 雌激素类药物引起子宫上皮细胞的过度增殖 (UTE)。孕酮(PG)因其抑制生长的作用而具有治疗作用。 转化生长因子-β介导的生长抑制由两个协同作用的受体转导 (Ri，RII)和下游的信号/转录因子Smad2/3， 激活阻止细胞周期进程的基因，如细胞周期蛋白依赖性 我们已经证明UTE从所有级别的UTE中分离出来 Eca通过诱导多种缺陷来逃避转化生长因子-β的负生长控制 转化生长因子-β的反应途径包括：转化生长因子-βRH的缺失、激活的Smad2和 P27kip1。此外，在ECA的前驱--复杂性增生(CH)中，这些 蛋白质已经减少了。因此，转化生长因子-b的作用很早就中断了。 在子宫内膜癌的发生中，提供了一个了解分子的机会 导致经济增长失调的事件。我们将使用正常的原代培养物， 和ECA细胞，并与基质细胞(UT)共培养，这与ECA细胞不同 品系中，保留了许多体内分化的特点。具体目标1， 将确定导致转化生长因子-b受体下调的分子机制 (例如，转录、翻译)和检测有缺陷的Smad2/3信号 使用转化生长因子-b启动子反应性报告试验。我们将努力恢复转化生长因子-β 瞬时将RII基因导入Eca UTE的功能。具体目标2 将检验Eca中p27kip1的丢失是通过以下途径降解的假设 泛素-蛋白酶体途径，它直接通过MAPK被E2-dnven所激活 通过RAS/MAPK/ERK1途径，而转化生长因子-b通常可以阻止p27的降解。 组织/腹水解物、蛋白酶体和MAPK的抑制剂以及免疫分析 将使用各种技术。使用单细胞和共培养，我们证明了UT 正常而非恶性子宫内膜介导PG诱导的生长抑制 正常的UTE。具体目标3将检验UTS的假设 旁分泌介导PG通过释放转化生长因子-β抑制UTE的生长 对PG的响应。我们将使用正常和“恶性”UT和UTE的混合培养 和由两者的UT组成的新型体内嵌合组织重组体，PG 受体敲除(PRKO)和野生型小鼠和人类UTE，这是荷尔蒙 在裸鼠和UTE体内移植，然后分析生长情况。这些 研究应阐明子宫内膜癌发生的分子机制， 激素(Dys)对子宫内膜生长的调节，并确定靶点 预防和治疗干预。