Departamento de Biología Celular y Genética, Universidad de Alcalá, Madrid, Spain.
Prostate. 2012 Jan;72(1):40-50. doi: 10.1002/pros.21404. Epub 2011 Apr 25.
The involvement of TNF-α in cancer development is controversial, since this cytokine was reported to act either as tumor promoter or suppressor. TNF-α may activate signaling pathways critical for life/death decisions, such as mitogen-activated protein kinases (MAPKs) and the anti-apoptotic NF-κB pathway. In this work, we investigate the activation status of NF-κB-related proteins in human prostate cancerous versus normal epithelium, and the alterations in the NF-κB pathway in relation to cell death in TNF-α-treated LNCaP (androgen-independent cells) and PC3 (androgen-independent) prostate cancer cell lines.
The expression of phospho-p38-MAPK, phospho-IKK-α/β and phospho-IκB-α, total IκB-α, and p65- and p50-NF-κB, were analyzed by immunohistochemistry in cancerous and normal prostate samples. The toxicity of TNF-α in LNCaP and PC3 cells, with or without kinase and NF-κB inhibitors, was assessed by changes on viability (MTT assay) and apoptosis (loss of DNA, annexin-V binding, and caspase cleavage/activation). Expression of NF-κB-related proteins in these cell lines was measured by Western blot.
Phospho-IκB-α, phospho-IKK-α/β and phospho-p38 levels, cytoplasmic p50 to IκB-α ratio, and nuclear p50 and p65, levels, were increased in cancerous epithelium, suggesting activation of the NF-κB pathway in prostatic malignance. TNF-α caused apoptosis with higher efficacy in LNCaP cells, and this response was potentiated by p38-MAPK inhibitor (LNCaP cells) and IKK-β inhibitor (both cell lines). However, the protective action of IKK-β was mediated by NF-κB only in LNCaP cells.
IKK-β mediates both NF-κB-dependent and -independent anti-apoptotic functions in prostate cancerous epithelium. IKK-β and p38-MAPK may represent useful therapeutic targets against prostate cancer.
TNF-α 参与癌症的发生发展的机制仍存在争议，因为这种细胞因子既可以作为肿瘤促进剂，也可以作为肿瘤抑制剂。TNF-α 可能激活对生死决策至关重要的信号通路，如丝裂原活化蛋白激酶（MAPKs）和抗细胞凋亡的 NF-κB 通路。在这项工作中，我们研究了人前列腺癌组织与正常上皮组织中 NF-κB 相关蛋白的激活状态，以及 TNF-α 处理的 LNCaP（雄激素非依赖性细胞）和 PC3（雄激素非依赖性）前列腺癌细胞系中与细胞死亡相关的 NF-κB 通路的变化。
通过免疫组织化学方法分析磷酸化 p38-MAPK、磷酸化 IKK-α/β 和磷酸化 IκB-α、总 IκB-α 以及 p65 和 p50-NF-κB 在癌组织和正常前列腺组织中的表达。用 MTT 法和 Annexin-V 结合实验及 caspase 切割/激活实验检测 TNF-α 在 LNCaP 和 PC3 细胞中的毒性作用，以及有无激酶和 NF-κB 抑制剂时的细胞活力。用 Western blot 检测这些细胞系中 NF-κB 相关蛋白的表达。
磷酸化 IκB-α、磷酸化 IKK-α/β 和磷酸化 p38 的水平、细胞质 p50 与 IκB-α 的比值、核内 p50 和 p65 的水平，在癌组织上皮中均增加，提示 NF-κB 通路在前列腺恶性肿瘤中被激活。TNF-α 在 LNCaP 细胞中诱导凋亡的作用更强，而 p38-MAPK 抑制剂（LNCaP 细胞）和 IKK-β 抑制剂（两种细胞系）则增强了这种反应。然而，只有在 LNCaP 细胞中，IKK-β 的保护作用才是通过 NF-κB 介导的。
IKK-β 在前列腺癌组织中具有 NF-κB 依赖性和非依赖性的抗凋亡功能。IKK-β 和 p38-MAPK 可能成为治疗前列腺癌的有用靶点。
