Increased L-arginine transport in a nitric oxide-producing metastatic colon cancer cell line
Increased L-arginine transport in a nitric oxide-producing metastatic colon cancer cell line
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DOI：
10.1007/bf02305770
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发表时间：
1996-09-01
影响因子：
3.7
通讯作者：
Lind, DS
中科院分区：
文献类型：
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作者：
Cendan, JC;Souba, WW;Lind, DS
背景：对人类肿瘤细胞中氨基酸转运的研究知之甚少。我们先前描述了人结肠癌细胞系SW 480中L-精氨酸的转运，发现它主要由钠非依赖性系统y（+）介导。本研究主要研究了L-精氨酸在转移细胞系SW 620中的转运，并与原代细胞系SW 480进行了比较。方法：在有和无钠的情况下，分析了H-3-L-精氨酸在细胞单层中的转运。在L-精氨酸浓度范围内进行动力学研究，以确定转运蛋白亲和力（Km）和最大转运速度（V-max）。通过用已知氨基酸阻断进一步表征转运。此外，细胞年龄的影响（即，培养时间）对精氨酸转运的影响。结果：SW 620细胞对L-精氨酸的摄取基本上不依赖于钠离子浓度，而对L-精氨酸的摄取则依赖于钠离子浓度。动力学和氨基酸抑制研究揭示了一个单一的高亲和力，钠非依赖性L-精氨酸转运蛋白（Vmax = 1286.3 +/- 158.3 pmol/mg蛋白/30 s; Km = 46.8 +/- 4.2 μ M）。钠非依赖性转运被系统y（+）底物L-高精氨酸、L-鸟氨酸和L-赖氨酸阻断，钠依赖性摄取通过具有系统B-O、B-+特征的单一转运蛋白发生（Km = 16.15 +/- 2.1 μ M; V-max = 329.94 +/- 29.7 pmol/mg蛋白/30 s）。精氨酸转运在培养中随时间增加，第2天细胞转运速度= 241.7 +/- 33.6 pmol/mg蛋白质/30 s，而第9天细胞转运速度= 377 +/- 15.4 pmol/mg蛋白质/30 s（p < 0.01）。细胞增殖研究显示，SW 620和SW 480的倍增时间分别为3.2天和5.4天（p < 0.05）。结论：在这些肿瘤细胞系中，L-精氨酸转运主要通过钠非依赖性、高亲和力的y（+）系统发生，在转移性变体（SW 620）中，Vmax增加了180%，表明y（+）转运蛋白上调。增加的y（+）活性可能是为肿瘤生长提供连续底物的机制。
Background: Little is known about amino acid transport in human neoplastic cells. We previously characterized L-arginine transport in the primary human colon cancer cell line, SW480, and found it is principally mediated by the sodium-independent system y(+). In this study, we characterized L-arginine transport in the metastatic cell line, SW620, and compared it with that in the primary cell line, SW480.Methods: Transport of H-3-L-arginine in cell monolayers was analyzed in the presence and absence of sodium. Kinetic studies were performed over a range of L-arginine concentrations to determine transporter affinity (K-m) and maximal transport velocity (V-max). Transport was further characterized through blockade with known amino acids. In addition, the effect of cell age (i.e., time in culture) on arginine transport was examined at 2 and 9 days after seeding. Cellular proliferation was asssessed by using the colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay.Results: L-Arginine uptake was primarily sodium independent in the SW620 cell line. Kinetic and amino acid-inhibition studies revealed a single high-affinity, sodium-independent L-arginine transporter (V-max = 1286.3 +/- 158.3 pmol/mg protein/30 s; K-m = 46.8 +/- 4.2 mu M). Sodium-independent transport was blocked by system y(+) substrates L-homoarginine, L-ornithine and L-lysine, Sodium-dependent uptake occurs through a single transporter with system B-O,B-+ characteristics (K-m = 16.15 +/- 2.1 mu M; V-max = 329.94 +/- 29.7 pmol/mg protein/30 s). Arginine transport increased with time in culture with day 2 cells transport velocity = 241.7 +/- 33.6 pmol/mg protein/30s, whereas day 9 cells transport velocity = 377 +/- 15.4 pmol/mg protein/30 s (p < 0.01). Cellular-proliferation studies revealed a doubling time of 3.2 days for SW620 and 5.4 days for SW480 (p < 0.05).Conclusions: L-Arginine transport in these neoplastic cell lines occurs primarily through sodium-independent, high-affinity system y(+), V-max was increased 180% in the metastatic variant (SW620), suggesting upregulation of the y(+) transporter. The increased y(+) activity may be a mechanism to provide continuous substrate for tumor growth.