Localization of the Na+-sugar cotransport system in a kidney epithelial cell line (LLC PK1).
Localization of the Na+-sugar cotransport system in a kidney epithelial cell line (LLC PK1).
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Na-糖共转运系统在肾上皮细胞系（LLC PK1）中的定位。
DOI：
10.1016/0005-2736(81)90417-x
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发表时间：
1981
期刊：
影响因子：
--
通讯作者：
Rabito,CA
中科院分区：
文献类型：
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作者：
Rabito,CA
对LLC PK 1细胞单层Na+依赖性糖转运定位的研究表明，甲基α-d-葡萄糖苷（一种共享葡萄糖-半乳糖转运系统的非代谢糖）的摄取主要发生在单层的顶侧。[3 H]根皮苷结合LLC PK 1细胞单层的动力学也进行了测量。这些研究表明存在两类不同的受体位点。高亲和力结合位点的解离常数（Kd）为1.2 μM，高亲和力受体的浓度为0.30 μmol/g DNA。另一类低亲和力位点的Kd为240 μM，结合位点数为12 μmol/g DNA。根皮苷在高亲和力结合位点的结合是Na+依赖性过程。相反，在低亲和力位点的结合是Na+非依赖性的。Na+对高亲和力结合位点的作用方式是增加解离常数而不改变结合位点的数目。根皮苷对甲基α-d-葡萄糖苷的抑制作用具有Na+依赖性，且高亲和力结合位点的Kd与根皮苷抑制甲基α-d-葡萄糖苷的Ki相匹配，表明根皮苷的高亲和力结合位点是甲基α-d-葡萄糖苷转运系统的一部分。从任何一方的单层的结合研究也表明，根皮苷在Na+依赖性高亲和力结合位点的结合主要发生在顶端，而不是基底侧。因此，LLC PK 1细胞顶端膜中Na+依赖性糖转运系统的特定位置是在组织培养条件下保留的上皮细胞功能极化的另一种表达。此外，由于这种糖转运在细胞悬浮后几乎消失，因此可以将其用作研究该细胞系中顶膜发育的标记。
Studies of the localization of the Na+-dependent sugar transport in monolayers of LLC PK 1 cells show that the uptake of a methyl α-d-glucoside, a nonmetabolizable sugar which shares the glucose-galactose transport system, occurs mainly from the apical side of the monolayer. Kinetics of [3 H] phlorizin binding to monolayers of LLC PK 1 cells were also measured. These studies demonstrate the presence of two distinct classes of receptor sites. The class comprising high affinity binding sites had a dissociation constant (K d) of 1.2 μM and a concentration of high affinity receptors of 0.30 μmol binding sites per g DNA. The other class involving low affinity sites had a K d of 240 μM with the number of binding sites equal to 12 μmol/g DNA. Phlorizin binding at high affinity binding sites is a Na+-dependent process. Binding at the low affinity sites on the contrary is Na+-independent. The mode of action of Na+ on the high affinity binding sites was to increase the dissociation constant without modifying the number of binding sites. The Na+ dependence and the matching of K d for high affinity binding sites with the K i of phlorizin for the inhibition of methyl α-d-glucoside strongly suggest that the high affinity phlorizin binding site is, or is part of the methyl α-d-glucoside transport system. Binding studies from either side of the monolayer also show that the binding of phlorizin at the Na+ dependent high affinity binding sites occurs mainly from the apical rather than the basolateral side. The specific location of the Na+-dependent sugar transport system in the apical membrane of LLC PK 1 cells is, therefore, another expression of the functional polarization of epithelial cells that is retained under tissue culture condition. In addition, since this sugar transport almost disappears after the cells are brought into suspension, it can be used as a marker to study the development of the apical membrane in this cell line.