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Involvement of PTEN in hormone-dependent proliferation of endometrial ocarcinoma cells

PTEN参与子宫内膜癌细胞激素依赖性增殖

基本信息

批准号：
14571552
负责人：
YOKOYAMA Yasuhiro
金额：
$ 2.24万
依托单位：
Gifu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

The fact that the genetic alterations of PTEN are frequently found in hormone-dependent cancers such as endometrial, breast, and prostata cancers, might suggest the involvement of PTEN in the hormone dependent cell growth of such tumors. Estrogen and estrogen-induced peptide growth factors promote the cell growth of endometrial glandular cells and would be associated with endometrial carcinogenesis. Endometrial carcinoma cells often lose steroid hormone-responsiveness during the carcinogenesis. We investigated whether PTEN modulates estrogen receptor status, the levels of enzymes of endogenous estrogen synthesis, and responsiveness against peptide growth factors in endometrial carcinoma cells. [Materials and methods] Recombinant adenovirus was produced by Adeno-X expression system. PTEN-null Ishikawa cell, an endometrial carcinoma cell line, was infected with the recombinant adenovirus at 20MOI to express efficiently PTEN protein. In Ishikawa cells expressing PTEN protein (PTEN-IK cell … More s), the followings were analyzed. 1)The expression levels of the enzymes for estrogen synthesis such as estrone sulfatase, estrone sulfotransferase, aromatase, 17 beta-hydroxysteroid dehydrogenases were analyzed by R-PCR. 2)The expression levels of estrogen receptors (alpha and beta) and their phosphorylated forms were analyzed by western blot and/or immunoprecipitation. 3)responsiveness against peptide growth factors including EGF was analyzed by MTT assay and western blot. [Results] (1)In-PTEN-IK cells, the estrone sulfatase and 17beta-hydroxysteroid dehydrogenase type 1 were upregulated when compared with those of parental Ishikawa cells. (2)The expression level of ER-beta was upregulated in PTEN-IK cells, but ER-alpha was not changed regardless of the downregutation of its phosphorylated form. (3)The cell growth of parental IK cells was significantly stimulated by EGF and IGF-1, and PTEN-IK cells were further sensitized the EGF or IGF1-growth stimulation. EFGR antibody could completely compromise the stimulatory effects of EGF in both cell lines. Wortmannin, a PI3K inhibitor, or UO126, a MPAK inhibitor, suppressed EGF-mediated cell growth stimulation partly in both cell lines. EGF augmented the level of phospho-Akt/PKB of PTEN-IK cells more effectively than that of parental IK cells. [Conclusion]PTEN regulates the level of ER-beta, phosphorylation level of ER-alpha, and intracellular estrogen concentration by modulating the levels of estrogen synthetic enzymes. Thus, the deficiency of PTEN protein alters estrogenic environment of the cells. Furthermore, the dysfunction of PTEN reduces responsiveness of EGF or other peptide growth factors of the cells. Endometrial carcinoma cells with deficient PTEN, thus, escape from hormone-dependency. Less

事实上，PTEN的基因改变经常发现在激素依赖性癌症，如子宫内膜癌，乳腺癌，前列腺癌，可能表明，PTEN参与激素依赖性细胞生长的肿瘤。雌激素及雌激素诱导肽类生长因子促进子宫内膜腺细胞生长，可能与子宫内膜癌的发生有关。子宫内膜癌细胞在癌变过程中常失去对激素的敏感性。我们研究了子宫内膜癌细胞中PTEN是否调节雌激素受体状态、内源性雌激素合成酶的水平以及对肽生长因子的反应性。[材料与方法]采用Adeno-X表达系统制备重组腺病毒。将重组腺病毒以20 MOI感染子宫内膜癌细胞系石川（Ishikawa），使其高效表达PTEN蛋白。在表达PTEN蛋白的石川细胞（PTEN-IK细胞）中， ...更多信息 s），进行以下分析。1)通过R-PCR分析雌激素合成酶如雌酮硫酸酯酶、雌酮磺基转移酶、芳香化酶、17 β-羟基类固醇脱氢酶的表达水平。2)通过蛋白质印迹和/或免疫沉淀分析雌激素受体（α和β）及其磷酸化形式的表达水平。3)用MTT法和Western blot法分析对包括EGF在内的肽类生长因子的反应性。[结果]（1）与亲本石川细胞相比，在PTEN-IK细胞中雌酮硫酸酯酶和17 β-羟基类固醇脱氢酶1型表达上调。（2）PTEN-IK细胞中ER-β表达上调，而ER-α磷酸化水平下调，ER-α表达无明显变化。（3）EGF和IGF-1对IK细胞的生长有明显的刺激作用，而PTEN-IK细胞对EGF和IGF-1的刺激作用更为敏感。EFGR抗体可完全破坏EGF在两种细胞系中的刺激作用。PI 3 K抑制剂Wortmannin或MPAK抑制剂UO 126在两种细胞系中部分抑制了EGF介导的细胞生长刺激。EGF能更有效地增加PTEN-IK细胞磷酸化Akt/PKB的表达。[结论]PTEN通过调节雌激素合成酶的水平，调节ER-β水平、ER-α磷酸化水平和细胞内雌激素浓度。因此，PTEN蛋白的缺乏改变了细胞的雌激素环境。此外，PTEN的功能障碍降低了细胞对EGF或其他肽生长因子的反应性。因此，具有缺陷的PTEN的子宫内膜癌细胞逃避了肿瘤依赖性。少