中文仓鼠卵巢（CHO-K1）细胞中酪氨酸酯对 125I-3-碘-α-甲基-L-酪氨酸摄取的刺激作用。
Stimulation of 125I-3-iodo-alpha-methyl-L-tyrosine uptake in Chinese hamster ovary (CHO-K1) cells by tyrosine esters.
机构信息
Department of Radiological Sciences, Center for Medical Sciences and Center for Humanities and Sciences, Ibaraki Prefectural University of Health Sciences, Inashiki-gun, Ibaraki, Japan.
出版信息
Nucl Med Biol. 2010 Feb;37(2):189-96. doi: 10.1016/j.nucmedbio.2009.10.003. Epub 2009 Nov 3.
INTRODUCTION
Transport of the amino acid analog (123)I-3-iodo-alpha-methyl-L-tyrosine, which is used in clinical SPECT imaging, occurs mainly via L-type amino acid transporter type 1 (LAT1; an amino acid exchanger). As LAT1 is highly expressed in actively proliferating tumors, we made a preliminary investigation of the effects of amino acid esters on enhancement of (125)I-3-iodo-alpha-methyl-L-tyrosine (IMT) uptake via LAT1 in Chinese hamster ovary (CHO-K1) cells.
METHODS
Because the sequence of the CHO-K1 LAT1 gene is not available, we confirmed LAT1 expression through IMT (18.5 kBq) uptake mechanisms using specific inhibitors. L-Gly, L-Ser, L-Leu, L-Phe, L-Met, L-Tyr, D-Tyr, L-Val and L-Lys ethyl/methyl esters were tested in combination with IMT. Time-course studies over a 3-h period were conducted, and the concentration dependence of L-Tyr ethyl and methyl esters (0.001 to 10 mM) in combination with IMT was also examined. For a proof of de-esterification of L- and D-Tyr ethyl and methyl esters in the cells (by enzymatic attack or other cause), the concentration of L- and D-Tyr was analyzed by high-performance liquid chromatography of the esters in phosphate buffer (pH 7.4) and cell homogenates at 37 degrees C or under ice-cold conditions.
RESULTS
Inhibition tests suggested that LAT1 is involved in IMT uptake by CHO-K1 cells. Co-administration of 1 mM of l-Tyr ethyl or methyl ester with IMT produced the greatest enhancement. The de-esterification reaction was stereo selective and temperature dependent in the homogenate. De-esterification kinetics were very fast in the homogenate and very slow in the phosphate buffer.
CONCLUSIONS
The L-Tyr ethyl or methyl esters were the most effective enhancers of IMT uptake into CHO-K1 cells and acted by trans-stimulation of the amino acid exchange function of LAT1. This result suggests that de-esterification in the cells may be caused by enzymatic attack. We will use IMT and L-Tyr ethyl or methyl esters to examine LAT1 function in tumor cells or tissues in vivo.
简介
用于临床 SPECT 成像的氨基酸类似物（123）I-3-碘-α-甲基-L-酪氨酸的转运主要通过 L 型氨基酸转运体 1（LAT1；一种氨基酸交换器）进行。由于 LAT1 在活跃增殖的肿瘤中高度表达，我们初步研究了氨基酸酯对中国仓鼠卵巢（CHO-K1）细胞中 LAT1 增强（125）I-3-碘-α-甲基-L-酪氨酸（IMT）摄取的影响。
方法
由于 CHO-K1 LAT1 基因的序列不可用，我们通过使用特定抑制剂的 IMT（18.5 kBq）摄取机制来确认 LAT1 表达。测试了 L-甘氨酸、L-丝氨酸、L-亮氨酸、L-苯丙氨酸、L-甲硫氨酸、L-酪氨酸、D-酪氨酸、L-缬氨酸和 L-赖氨酸乙酯/甲酯与 IMT 的组合。进行了 3 小时的时间过程研究，并检查了 L-酪氨酸乙酯和甲酯（0.001 至 10 mM）与 IMT 组合的浓度依赖性。为了证明 L-和 D-酪氨酸乙酯和甲酯在细胞中的（通过酶攻击或其他原因）去酯化，在 37°C 或冰冷条件下，通过高效液相色谱法分析磷酸盐缓冲液（pH 7.4）和细胞匀浆中的 L-和 D-酪氨酸的浓度。
结果
抑制试验表明 LAT1 参与 CHO-K1 细胞中 IMT 的摄取。与 IMT 一起给予 1 mM 的 l-酪氨酸乙酯或甲酯可产生最大的增强效果。在匀浆中，去酯化反应具有立体选择性和温度依赖性。在匀浆中，去酯化动力学非常快，而在磷酸盐缓冲液中非常慢。
结论
L-酪氨酸乙酯或甲酯是增强 CHO-K1 细胞摄取 IMT 的最有效增强剂，通过 LAT1 的氨基酸交换功能的反式刺激起作用。这一结果表明，细胞中的去酯化可能是由酶攻击引起的。我们将使用 IMT 和 L-酪氨酸乙酯或甲酯在体内检查肿瘤细胞或组织中的 LAT1 功能。
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