Induction of G0/G1 cell cycle arrest in ovarian carcinoma cells by the anti-inflammatory drug NS-398, but not by COX-2-specific RNA interference
Induction of G0/G1 cell cycle arrest in ovarian carcinoma cells by the anti-inflammatory drug NS-398, but not by COX-2-specific RNA interference
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DOI：
10.1038/sj.onc.1206920
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发表时间：
2003-11-27
期刊：
影响因子：
8
通讯作者：
Hauptmann, S
中科院分区：
文献类型：
--
作者：
Denkert, C;Fürstenberg, A;Hauptmann, S
环氧合酶，特别是考克斯-2，在肿瘤的发生和发展中起重要作用。我们以前的研究表明，考克斯-2的表达是一个独立的预后因素，在人类卵巢癌。在这项研究中，我们研究了抑制考克斯亚型的NSAID NS-398以及COX亚型特异性RNA干扰（RNAi）在人卵巢癌细胞系OVCAR-3和SKOV-3的影响。OVCAR-3细胞表现出考克斯-1的组成性表达，并在白细胞介素-1 β刺激后诱导高水平的考克斯-2和PGE（2）。相反，SKOV-3细胞对两种考克斯亚型均为阴性。在OVCAR-3细胞中，1 μ M浓度的NS-398和考克斯-2特异性沉默RNA（siRNA）抑制PGE（2）的产生，而考克斯-1特异性siRNA则没有影响。这表明，考克斯-2是该细胞系中PGE（2）的主要来源。为了区分考克斯-2特异性和非考克斯-2特异性对细胞增殖的影响，在用NS-398和考克斯siRNA孵育细胞后进行增殖测定。NS-398在50-500 μ M的浓度下诱导细胞增殖的抑制，该浓度高于抑制PGE（2）产生所需的浓度。这种抑制作用存在于COX阳性细胞系OVCAR-3和COX阴性细胞系SKOV-3中，并且不能通过添加外源性PGE而逆转（2）。考克斯-1和考克斯-2特异性siRNA均不影响OVCAR-3细胞的细胞增殖。细胞周期分析显示，用NS-398处理后，G 0/G1期细胞的积累增加，但用考克斯siRNA处理后没有。这些实验表明，NS-398通过诱导G 0/G1细胞周期阻滞而降低卵巢癌细胞的细胞增殖，而不依赖于考克斯-2抑制。我们的研究表明，通过RNAi特异性抑制考克斯亚型可用于分离NSAID的作用。此外，我们的研究结果表明，细胞周期阻滞是负责NS-398对卵巢癌细胞的抗增殖作用的主要机制之一。
Cyclooxygenases, particularly COX-2, play an important role in tumor development and progression. We have previously shown that COX-2 expression is an independent prognostic factor in human ovarian carcinoma. In this study, we investigated the effects of the inhibition of COX isoforms by the NSAID NS-398 as well as by COX-isoform-specific RNA interference (RNAi) in the human ovarian carcinoma cell lines OVCAR-3 and SKOV-3. OVCAR-3 cells showed a constitutive expression of COX-1 and an induction of high levels of COX-2 and PGE(2) after stimulation with interleukin-1beta. In contrast, SKOV-3 cells were negative for both COX isoforms. In OVCAR-3 cells, PGE(2) production was inhibited by NS-398 in concentrations of 1 muM and by a COX-2-specific silencing RNA ( siRNA), while a COX-1-specific siRNA did not have an effect. This suggests that COX-2 is the major source of PGE(2) in this cell line. To dissociate COX-2-specific and non-COX-2-specific effects on cell proliferation, a proliferation assay was performed after incubation of cells with NS-398 and COX siRNAs. NS-398 induced an inhibition of cell proliferation at concentrations of 50-500 muM, which are above the concentrations needed for the inhibition of PGE(2) production. This inhibitory effect was present in the COX-positive cell line OVCAR-3 as well as in the COX-negative cell line SKOV-3 and could not be reverted by addition of exogenous PGE(2). Neither COX-1- nor COX-2-specific siRNAs had an effect on cell proliferation of OVCAR-3 cells. Cell cycle analysis showed an increased accumulation of cells in the G0/G1 phase after treatment with NS-398, but not with COX siRNAs. These experiments suggest that NS-398 reduced cell proliferation in ovarian carcinoma cells by induction of G0/G1 cell cycle arrest independent of COX-2 inhibition. Our study shows that specific inhibition of COX isoforms by RNAi could be used to dissociate effects of NSAIDs. Furthermore, our results suggest that cell cycle arrest is one of the primary mechanisms responsible for the antiproliferative effects of NS-398 on ovarian carcinoma cells.