Mechanisms and regulation of vitamin C uptake: studies of the hSVCT systems in human liver epithelial cells
Mechanisms and regulation of vitamin C uptake: studies of the hSVCT systems in human liver epithelial cells
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DOI：
10.1152/ajpgi.90399.2008
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发表时间：
2008-12-01
影响因子：
4.5
通讯作者：
Said, Hamid M.
中科院分区：
文献类型：
--
作者：
Reidling, Jack C.;Subramanian, Veedamali S.;Said, Hamid M.
人类使用两种钠-抗坏血酸共转运体(hSVCT1和hSVCT2)来运输膳食中必需的微量营养素抗坏血酸，抗坏血酸是维生素C的还原和活性形式。虽然人体肝脏在调节和维持维生素C动态平衡方面起着关键作用，但维生素C的转运生理学和hSVCT系统在该器官中的调节还没有很好的定义。因此，这项研究使用了人肝细胞系(HepG2)，用原代人肝细胞证实了某些结果，并确定了抗坏血酸的初始摄取速度是Na(+)梯度的，依赖于pH，在低和高微摩尔范围内是饱和的浓度函数。此外，hSVCT2蛋白和mRNA在HepG2细胞和人肝组织中均有较高水平的表达，并且克隆的hSVCT2启动子在HepG2细胞中具有更高的活性。使用短干扰RNA的结果表明，在HepG2细胞中，降低hSVCT2消息水平比降低hSVCT1消息水平更能减少整体抗坏血酸摄取过程。激活PKC细胞内调节通路导致抗坏血酸摄取的下调，而不是由hSVCT1或hSVCT2中单个预测的PKC特异性氨基酸磷酸化位点介导的。然而，PKC的激活导致hSVCT1的内化，而不是hSVCT2的内化。对其他细胞内抗坏血酸摄取调节通路的研究表明，PKA、PTK和Ca(2+)/钙调蛋白也可能调节抗坏血酸摄取，但不是通过一氧化氮依赖的途径。这些研究首次确定了人肝上皮细胞摄取抗坏血酸的整个过程以及hSVCT系统的相对表达、调节和贡献。
Humans use two sodium-ascorbate cotransporters (hSVCT1 and hSVCT2) for transporting the dietary essential micronutrient ascorbic acid, the reduced and active form of vitamin C. Although the human liver plays a pivotal role in regulating and maintaining vitamin C homeostasis, vitamin C transport physiology and regulation of the hSVCT systems in this organ have not been well defined. Thus, this research used a human hepatic cell line (HepG2), confirming certain results with primary human hepatocytes and determined the initial rate of ascorbic acid uptake to be Na(+) gradient, pH dependent, and saturable as a function of concentration over low and high micromolar ranges. Additionally, hSVCT2 protein and mRNA are expressed at higher levels in HepG2 cells and native human liver, and the cloned hSVCT2 promoter has more activity in HepG2 cells. Results using short interfering RNA suggest that in HepG2 cells, decreasing hSVCT2 message levels reduces the overall ascorbic acid uptake process more than decreasing hSVCT1 message levels. Activation of PKC intracellular regulatory pathways caused a downregulation in ascorbic acid uptake not mediated by a single predicted PKC-specific amino acid phosphorylation site in hSVCT1 or hSVCT2. However, PKC activation causes internalization of hSVCT1 but not hSVCT2. Examination of other intracellular regulatory pathways on ascorbic acid uptake determined that regulation also potentially occurs by PKA, PTK, and Ca(2+)/calmodulin, but not by nitric oxide-dependent pathways. These studies are the first to determine the overall ascorbic acid uptake process and relative expression, regulation, and contribution of the hSVCT systems in human liver epithelial cells.