Pyrrolidine dithiocarbamate exerts anti-proliferative and pro-apoptotic effects in renal cell carcinoma cell lines
Pyrrolidine dithiocarbamate exerts anti-proliferative and pro-apoptotic effects in renal cell carcinoma cell lines
复制标题
DOI：
10.1093/ndt/gfl543
复制
发表时间：
2006-12-01
影响因子：
6.1
通讯作者：
Healy, Helen
中科院分区：
文献类型：
--
作者：
Morais, Christudas;Pat, Betty;Healy, Helen
背景核因子-κ B（NF-κ B）的活化与肾细胞癌（RCC）的发生、发展和转移有关。本研究探讨了NF-κ B B抑制剂吡咯烷二硫代氨基甲酸酯（PDTC）对两种转移性肾癌细胞系ACHN和SN 12 K1的作用。将RCC细胞系和正常细胞暴露于25或50 μ M的PDTC。流式细胞术和TdT介导的缺口末端标记法检测细胞凋亡。MTT法检测细胞活力，BrdU法检测细胞增殖。通过免疫印迹和/或免疫荧光分析NF-κ B亚单位、I κ B、I κ B激酶（IKK）复合物和凋亡调节蛋白的表达。ELISA法检测NF-κ B B亚基的DNA结合活性。RCC细胞系的NF-κ B B的所有五个亚基的基础水平表达高于正常的人近端肾小管上皮细胞或HK-2细胞的原代培养物。PDTC可降低肾细胞癌细胞的存活率和增殖能力，但对正常细胞无明显影响。在两种RCC细胞系中，ACHN的所有五种NF-κ B亚基的基础水平表达均高于SN 12 K1，并且对PDTC更具抗性。虽然PDTC诱导两种RCC细胞系中所有五种NF-κ B亚基表达的总体降低，但出乎意料的是，它增加了ACHN中NF-κ B的核表达，但在SN 12 K1中没有。PDTC降低了所有NF-κ B亚基的DNA结合活性和IKK复合物（IKK-α、IKK-β和IKK-γ）以及抑制单位I κ B-α和I κ B-β的表达。PDTC诱导两种RCC细胞系凋亡的显着增加。这与抗凋亡蛋白Bcl-2和Bcl-（XL）的表达减少有关，而促凋亡蛋白Bcl-2的表达没有明显变化。这些数据表明，PDTC有可能成为某些形式的RCC的抗癌剂。
Background. The activation of nuclear factor-kappa B (NF-kappa B) has been implicated in the development, progression and metastasis of renal cell carcinoma (RCC). This study investigates the effect of pyrrolidine dithiocarbamate (PDTC), a NF-kappa B inhibitor, on two metastatic human RCC cell lines, ACHN and SN12K1.Methods. RCC cell lines and normal cells were exposed to 25 or 50 mu M of PDTC. Apoptosis was measured by flow cytometry and TdT-mediated nick end labelling methods. Cell viability and proliferation were measured by MTT and BrdU assays, respectively. Expression of NF-kappa B subunits, I kappa Bs, I kappa B Kinase (IKK) complex and apoptotic regulatory proteins were analysed by western blotting and/or immunofluorescence. DNA-binding activity of NF-kappa B subunits were measured by ELISA.Results. RCC cell lines had a higher basal level expression of all the five subunits of NF- kappa B than normal primary cultures of human proximal tubular epithelial cells or HK-2 cells. PDTC decreased the viability and proliferation of RCC, but not normal cells. Of the two RCC cell lines, ACHN had a higher basal level expression of all the five NF-kappa B subunits than SN12K1 and was more resistant to PDTC. While PDTC induced an overall decrease in expression of all the five NF-kappa B subunits in both RCC cell lines, unexpectedly, it increased the nuclear expression of NF-kappa B in ACHN, but not in SN12K1. PDTC reduced the DNA-binding activity of all the NF-kappa B subunits and the expression of the IKK complex (IKK-alpha, IKK-beta and IKK-gamma) and the inhibitory units I kappa B-alpha and I kappa B-beta. PDTC induced a significant increase in apoptosis in both RCC cell lines. This was associated with a decrease in expression of the anti-apoptotic proteins, Bcl-2 and Bcl-(XL), without marked changes in the pro-apoptotic protein Bax.Conclusion. These data suggest that PDTC has the potential to be an anticancer agent in some forms of RCC.