Institution of Biochemistry and Molecular Biology, Medical School of Shandong University, Jinan 250012, China.
Biochem Biophys Res Commun. 2010 Jul 30;398(3):457-61. doi: 10.1016/j.bbrc.2010.06.099. Epub 2010 Jul 1.
NKX3.1, a prostate-specific homeobox gene, plays an important role in prostate cancer and usually functions as tumor suppressor gene. Previously we have demonstrated that forced expression of NKX3.1 reduced cell growth and invasion in prostate cancer cell line PC-3. Presently, we investigated the effect of NKX3.1 on the sensitivity of the prostate cancer cells to apoptosis inducer tumor necrosis factor-alpha (TNF-alpha) and cycloheximide (CHX). PC-3 cells were transfected with NKX3.1 expression plasmid (pcDNA3.1-NKX3.1) and LNCaP cells were transfected with siRNA expression plasmid (pRNAT-RNAi1) targeting NKX3.1. The cell morphology and apoptotic rate were analyzed by Hoechst 33342 staining and Flow Cytometry in absence or presence of TNF-alpha and CHX. The activity of caspase-3 was determined using DEVD-pNA as substrate. Simultaneously, the effect of NKX3.1 on caspase-3 expression was detected using RT-PCR and Western blot. The results showed that ectopic expression of NKX3.1 promoted TNF-alpha/CHX-induced apoptosis in PC-3 cells, whereas knockdown of NKX3.1 protected LNCaP cells from apoptosis induced by TNF-alpha/CHX. The pro-apoptosis activity of NKX3.1 might partially contribute to its elevation of caspase-3 expression and activity. Manipulating NKX3.1 expression should be a promising therapeutic strategy for treating both androgen-dependent and androgen-independent prostate cancer.
NKX3.1 是一种前列腺特异性同源盒基因，在前列腺癌中发挥重要作用，通常作为肿瘤抑制基因发挥作用。先前我们已经证明，NKX3.1 的强制表达可降低前列腺癌细胞系 PC-3 的细胞生长和侵袭。目前，我们研究了 NKX3.1 对前列腺癌细胞对凋亡诱导剂肿瘤坏死因子-α（TNF-α）和环己酰亚胺（CHX）敏感性的影响。用 NKX3.1 表达质粒（pcDNA3.1-NKX3.1）转染 PC-3 细胞，用靶向 NKX3.1 的 siRNA 表达质粒（pRNAT-RNAi1）转染 LNCaP 细胞。在不存在或存在 TNF-α和 CHX 的情况下，通过 Hoechst 33342 染色和流式细胞术分析细胞形态和凋亡率。使用 DEVD-pNA 作为底物测定 caspase-3 的活性。同时，使用 RT-PCR 和 Western blot 检测 NKX3.1 对 caspase-3 表达的影响。结果表明，NKX3.1 的异位表达促进了 PC-3 细胞中 TNF-α/CHX 诱导的细胞凋亡，而 NKX3.1 的敲低则保护了 LNCaP 细胞免受 TNF-α/CHX 诱导的凋亡。NKX3.1 的促凋亡活性可能部分归因于其 caspase-3 表达和活性的提高。操纵 NKX3.1 的表达可能是治疗雄激素依赖性和雄激素非依赖性前列腺癌的一种很有前途的治疗策略。
