Expression profile and functional activity of peptide transporters in prostate cancer cells.
Expression profile and functional activity of peptide transporters in prostate cancer cells.
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DOI：
10.1021/mp300364k
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发表时间：
2013-02-04
影响因子：
4.9
通讯作者：
Cheng K
中科院分区：
文献类型：
--
作者：
Tai W;Chen Z;Cheng K
肽转运蛋白主要在肠和肾上皮细胞中表达。肽转运蛋白的功能性表达也在其他类型的组织中鉴定，例如神经胶质细胞、巨噬细胞和胆管、肺和乳腺的上皮细胞。然而，它们的存在和作用在癌症中知之甚少。我们探索了前列腺癌细胞系LNCaP、PC-3和DU 145中肽转运蛋白的表达谱和功能活性。采用定量真实的时间PCR（qRT-PCR）和蛋白质印迹法（Western blot）来评估这些细胞中肽转运蛋白1（PEPT 1）、肽转运蛋白2（PEPT 2）、肽组氨酸转运蛋白1（PHT 1）和肽组氨酸转运蛋白2（PHT 2）的表达谱。LNCaP表达高水平的PEPT 2和PHT 1，而PC-3显示PEPT 1和PHT 1的强表达。DU 145仅显示PEPT 1和PHT 1的弱表达。使用放射性标记的甘氨酰肌氨酸（[3 H]Gly-Sar）和L-组氨酸（[3 H] L-组氨酸）在这些细胞系中研究功能活性。[3 H]Gly-Sar和[3 H] L-组氨酸的摄取具有时间和pH依赖性。动力学研究表明，在检测浓度范围内，Gly-Sar和L-组氨酸的摄取可饱和。在三种细胞系中表现出的结合亲和力（Km）和最大速度（Vmax）与我们在qPCR和Western印迹分析中观察到的表达谱一致。竞争性抑制研究表明，前列腺癌细胞中的肽转运蛋白表现出广泛的底物特异性，偏好疏水性二肽，如Leu-Leu。荧光显微镜观察发现，肽转运蛋白底物D-Ala-Lys-AMCA在LNCaP和PC-3细胞胞质中特异性聚集，而在DU 145细胞胞质中不聚集。Gly-Sar抑制肽转运蛋白活性可抑制LNCaP和PC-3细胞的生长。本研究表明PC-3细胞可作为研究PEPT 1的一种新的细胞培养模型，LNCaP可作为研究PEPT 2的一种模型。此外，我们的研究结果表明，肽转运蛋白在前列腺癌细胞中过表达，可以作为一个有前途的目标，为肿瘤特异性药物输送。
Peptide transporters are expressed predominantly in intestinal and renal epithelial cells. The functional expression of peptide transporters is also identified in other types of tissues, such as glia cells, macrophages, and the epithelia of the bile duct, the lungs and the mammary glands. However, their presence and role are poorly understood in carcinomas. We explored the expression profile and functional activity of peptide transporters in the prostate cancer cell lines LNCaP, PC-3 and DU145. Quantitative Real Time PCR (qRT-PCR) and Western blot were used to evaluate the expression profile of peptide transporter 1 (PEPT1), peptide transporter 2 (PEPT2), peptide histidine transporter 1 (PHT1) and peptide histidine transporter 2 (PHT2) in these cells. LNCaP expresses high levels of PEPT2 and PHT1, while PC-3 demonstrates strong expression of PEPT1 and PHT1. DU145 shows only weak expression of PEPT1 and PHT1. Functional activities were studied in these cell lines using radiolabeled Glycylsarcosine ([3H]Gly-Sar) and L-Histidine ([3H]L-Histidine). The uptake of [3H]Gly-Sar and [3H]L-Histidine was time- and pH-dependent. A kinetic study showed that the uptake of Gly-Sar and L-Histidine is saturable over the tested concentration range. The binding affinity (Km) and the maximal velocity (Vmax) exhibited in the three cell lines were consistent with the expression profiles we observed in qPCR and Western blot analysis. A competitive inhibition study revealed that peptide transporters in prostate cancer cells exhibited broad substrate specificity with a preference for hydrophobic dipeptides, such as Leu-Leu. Fluorescence microscopy revealed that the fluorescent dipeptide probe D-Ala-Lys-AMCA (a substrate of peptide transporters) specifically accumulated in the cytoplasm of LNCaP and PC-3, but not DU145 cells. Inhibiting the peptide transporter activity by Gly-Sar suppressed the growth of LNCaP and PC-3 cells. Our study indicated that PC-3 cells can be established as a new cell culture model for PEPT1 study, and LNCaP can be used as a model for PEPT2 study. Moreover, our results suggested that peptide transporters are over-expressed in prostate cancer cells and can be adopted as a promising target for tumor-specific drug delivery.
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影响因子：
3.7
作者：
Qin, Bin;Tai, Wanyi;Shukla, Ravi S.;Cheng, Kun
通讯作者：
Cheng, Kun
影响因子：
6.4
作者：
Nawashiro, Hiroshi;Otani, Naoki;Endou, Hitoshi
通讯作者：
Endou, Hitoshi
DOI：
10.1016/j.ejpb.2004.07.008
发表时间：
2005-01-01
影响因子：
4.9
作者：
Luckner, P;Brandsch, M
通讯作者：
Brandsch, M
影响因子：
4.5
作者：
Mackenzie, B;Erickson, JD
通讯作者：
Erickson, JD
DOI：
10.1152/ajpgi.00534.2001
发表时间：
2002-07-01
影响因子：
4.5
作者：
Knütter, I;Rubio-Aliaga, I;Brandsch, M
通讯作者：
Brandsch, M