Sumitomo M, Tachibana M, Nakashima J, Murai M, Miyajima A, Kimura F, Hayakawa M, Nakamura H
Department of Urology, National Defense Medical College, Tokorozawa, Saitama, Japan.
J Urol. 1999 Feb;161(2):674-9.
Although tumor necrosis factor-alpha (TNF-alpha) induces a strong cytotoxic effect on cell growth, many authors have reported that various cancer cells are resistant to TNF-alpha and the basis for this sensitivity or resistance to TNF-alpha remains to be elucidated. Since nuclear factor kappa B (NF-kappaB) activation has recently been reported to inhibit TNF-alpha-induced cell death, we studied whether NF-kappaB also assumes a protective role in TNF-alpha-induced cell death in prostate cancer cells.
We used two human prostate cancer cell lines of DU145 and PC-3. We prepared two different NF-kappaB inhibitors, pyrrolidine dithiocarbamate (PDTC) and NF-kappaB decoy. NF-kappaB DNA binding activity was detected by electrophoretic mobility shift assay (EMSA). Cell survivals were measured by MTT assay. Induction of apoptosis was detected by nuclear staining and measured by fragmented DNA ELISA.
EMSA showed that NF-kappaB inhibitors continuously inhibited TNF-alpha-induced NF-kappaB activation. Cell growth was not inhibited by either TNF-alpha (50 ng./ml. or less) or NF-kappaB inhibitors. However, both PCA cells treated with TNF-alpha (20 ng./ml.) plus NF-kappaB inhibitors showed significant growth inhibition compared with controls (p<0.05). Nuclei of PCA cells appeared severely fragmented by this combination therapy. Furthermore, the levels of DNA fragmentation were significantly elevated in PCA cells treated with TNF-alpha (20 ng./ml.) plus NF-kappaB inhibitors compared with controls (p<0.05).
NF-kappaB activation is suggested to produce the resistance of DU145 and PC-3 to TNF-alpha and that the combination of TNF-alpha and NF-kappaB inhibitors could be constituted an effective therapy to TNF-alpha-resistant human prostate cancer cells.
尽管肿瘤坏死因子-α（TNF-α）对细胞生长具有强大的细胞毒性作用，但许多作者报告称，各种癌细胞对TNF-α具有抗性，而这种对TNF-α敏感或抗性的基础仍有待阐明。由于最近有报道称核因子κB（NF-κB）激活可抑制TNF-α诱导的细胞死亡，我们研究了NF-κB在TNF-α诱导的前列腺癌细胞死亡中是否也发挥保护作用。
我们使用了DU145和PC-3两种人前列腺癌细胞系。我们制备了两种不同的NF-κB抑制剂，吡咯烷二硫代氨基甲酸盐（PDTC）和NF-κB诱饵。通过电泳迁移率变动分析（EMSA）检测NF-κB DNA结合活性。通过MTT法测量细胞存活率。通过细胞核染色检测凋亡诱导情况，并通过DNA片段化ELISA进行测量。
EMSA显示NF-κB抑制剂持续抑制TNF-α诱导的NF-κB激活。TNF-α（50 ng/ml或更低）或NF-κB抑制剂均未抑制细胞生长。然而，与对照组相比，用TNF-α（20 ng/ml）加NF-κB抑制剂处理的两种前列腺癌细胞系均显示出显著的生长抑制（p<0.05）。这种联合治疗使前列腺癌细胞的细胞核出现严重碎片化。此外，与对照组相比，用TNF-α（20 ng/ml）加NF-κB抑制剂处理的前列腺癌细胞中DNA片段化水平显著升高（p<0.05）。
提示NF-κB激活使DU145和PC-3对TNF-α产生抗性，并且TNF-α与NF-κB抑制剂的联合应用可能构成对TNF-α抗性人前列腺癌细胞的有效治疗方法。
