Horie Asuka, Ishida Kazuya, Watanabe Yuri, Shibata Kaito, Hashimoto Yukiya
Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama, 930-0194, Japan.
Biopharm Drug Dispos. 2014 Dec;35(9):532-42. doi: 10.1002/bdd.1917. Epub 2014 Oct 29.
The aim of the present study was to investigate the membrane transport mechanisms of choline using human intestinal epithelial LS180 cells. The mRNA of choline transporter-like proteins (CTLs) was expressed significantly in LS180 cells, and the rank order was CTL1 > CTL4 > CTL3 > CTL2 > CTL5. In contrast, the mRNA expression of other choline transporters, organic cation transporter (OCT) 1, OCT2 and high-affinity choline transporter 1 (CHT1), was considerably lower in LS180 cells. Five mm unlabelled choline, hemicolinium-3 and guanidine, but not tetraethylammonium, inhibited the cellular uptake of 100 µm choline in LS180 cells. The uptake of choline into LS180 cells was virtually Na(+)-independent. The uptake of choline was significantly decreased by acidification of the extracellular pH; however, it was not increased by alkalization of the extracellular pH. In addition, both acidification and alkalization of intracellular pH decreased the uptake of choline, indicating that the choline uptake in LS180 cells is not stimulated by the outward H(+) gradient. On the other hand, the uptake of choline was decreased by membrane depolarization along with increasing extracellular K(+) concentration. In addition, the Na(+)-independent uptake of choline was saturable, and the Km value was estimated to be 108 µm. These findings suggest that the uptake of choline into LS180 cells is membrane potential-dependent, but not outward H(+) gradient-dependent.
本研究的目的是利用人肠上皮LS180细胞研究胆碱的膜转运机制。胆碱转运样蛋白（CTLs）的mRNA在LS180细胞中显著表达，其表达顺序为CTL1>CTL4>CTL3>CTL2>CTL5。相比之下，其他胆碱转运体，即有机阳离子转运体（OCT）1、OCT2和高亲和力胆碱转运体1（CHT1）的mRNA在LS180细胞中的表达则低得多。5 mM未标记的胆碱、半胱氨酸-3和胍，但不是四乙铵，可抑制LS180细胞对100 µm胆碱的细胞摄取。胆碱进入LS180细胞的摄取实际上不依赖于Na(+)。细胞外pH值酸化显著降低了胆碱的摄取；然而，细胞外pH值碱化并没有增加胆碱的摄取。此外，细胞内pH值的酸化和碱化均降低了胆碱的摄取，这表明LS180细胞中胆碱的摄取不受外向H(+)梯度的刺激。另一方面，随着细胞外K(+)浓度的增加，膜去极化降低了胆碱的摄取。此外，胆碱的不依赖Na(+)的摄取是可饱和的，Km值估计为108 µm。这些发现表明，胆碱进入LS180细胞的摄取是膜电位依赖性的，但不是外向H(+)梯度依赖性的。
