Berry Emily, Hardt Jennifer L, Clardy Jon, Lurain John R, Kim J Julie
Department of Obstetrics and Gynecology, Division of Gynecologic Oncology, Northwestern University, Chicago, IL 60611, USA.
Gynecol Oncol. 2009 Feb;112(2):331-6. doi: 10.1016/j.ygyno.2008.10.017. Epub 2008 Nov 28.
Endometrial cancer is the most common type of gynecologic cancer in the United States. In this study, we propose that a marine sponge compound, psammaplysene A (PsA) induces apoptosis in endometrial cancer cells through forced nuclear expression of FOXO1.
Ishikawa and ECC1 cells were treated with varying doses of PsA. FOXO1 protein localization was observed using immunofluorescent staining of cells. The effects of PsA on cell viability and proliferation were assessed using a cell viability assay and a BrdU incorporation assay respectively. Cell cycle analysis was performed using flow cytometry. To assess the role of FOXO1 in PsA-induced apoptosis, FOXO1 was silenced in ECC1 cells using siRNA technique, and overexpressed in Ishikawa cells using an adenovirus containing FOXO1 cDNAs. Western blots were used to measure levels of FOXO1 and cleaved PARP proteins.
Treatment of both ECC1 and Ishikawa cells with PsA caused an increase in nuclear FOXO1 protein, a dramatic decrease in cell viability of approximately 5-fold (p<0.05) and minimal effect on proliferation. Furthermore, treatment of cells with PsA doubled the percentage of cells in the G2/M phase (p<0.05). PsA induced apoptosis in endometrial cancer cells. When FOXO1 was silenced in ECC1 cells and treated with PsA, the incidence of apoptosis decreased. In addition, overexpression of FOXO1 with PsA treatment increased apoptosis.
Increasing nuclear FOXO1 function is important for the induction of apoptosis of endometrial cancer cells by PsA.
子宫内膜癌是美国最常见的妇科癌症类型。在本研究中，我们提出一种海洋海绵化合物——沙海葵毒素A（PsA）通过强制FOXO1在细胞核中表达来诱导子宫内膜癌细胞凋亡。
用不同剂量的PsA处理Ishikawa细胞和ECC1细胞。通过细胞免疫荧光染色观察FOXO1蛋白的定位。分别使用细胞活力测定法和BrdU掺入测定法评估PsA对细胞活力和增殖的影响。使用流式细胞术进行细胞周期分析。为了评估FOXO1在PsA诱导的凋亡中的作用，利用小干扰RNA（siRNA）技术使ECC1细胞中的FOXO1沉默，并利用含有FOXO1 cDNA的腺病毒使Ishikawa细胞中的FOXO1过表达。使用蛋白质免疫印迹法检测FOXO1和裂解的PARP蛋白的水平。
用PsA处理ECC1细胞和Ishikawa细胞均导致细胞核内FOXO1蛋白增加，细胞活力显著下降约5倍（p<0.05），对增殖的影响最小。此外，用PsA处理细胞使处于G2/M期的细胞百分比增加了一倍（p<0.05）。PsA诱导子宫内膜癌细胞凋亡。当ECC1细胞中的FOXO1沉默并用PsA处理时，凋亡发生率降低。此外，PsA处理下FOXO1过表达增加了细胞凋亡。
增强细胞核内FOXO1功能对于PsA诱导子宫内膜癌细胞凋亡至关重要。
