Bellovino D, Lanyau Y, Garaguso I, Amicone L, Cavallari C, Tripodi M, Gaetani S
Istituto Nazionale della Nutrizione, Rome, Italy.
J Cell Physiol. 1999 Oct;181(1):24-32. doi: 10.1002/(SICI)1097-4652(199910)181:1<24::AID-JCP3>3.0.CO;2-0.
The untransformed stable cell line Met murine hepatocytes (MMH), generated from liver explants of transgenic mice expressing a constitutively active truncated form of the human hepatocyte growth factor receptor (cyto-Met), represents an innovative tool for in vitro studies of liver function. In the present report, we show that the MMH-D3 line isolated from the liver of a 3-day-old mouse is a useful model to investigate the regulation of the synthesis and secretion of retinol-binding protein (RBP) by retinol (vitamin A alcohol). Experiments with Northern blot hybridization, metabolic labeling of cellular proteins followed by immunoprecipitation, and Western blot analysis demonstrated that, similarly to the in vivo situation, in MMH-D3 cells the presence of retinol does not affect transcriptional and translational rate of the RBP gene but is essential for regulating the secretion rate of the protein. Unlike HepG2 human hepatocarcinoma cells used thus far in studies of retinoid metabolism, including the synthesis and secretion of RBP, vitamin A deficiency causes, in MMH-D3 cells, the inhibiton of RBP secretion and the protein accumulation in the cell, whereas retinol repletion promptly results in RBP secretion. This model will be very useful in future studies on vitamin A distribution in the organism.
未转化的稳定细胞系Met小鼠肝细胞（MMH）由表达人肝细胞生长因子受体组成型活性截短形式（细胞Met）的转基因小鼠肝脏外植体产生，是肝功能体外研究的一种创新工具。在本报告中，我们表明从3日龄小鼠肝脏分离的MMH-D3细胞系是研究视黄醇（维生素A醇）对视黄醇结合蛋白（RBP）合成和分泌调节的有用模型。通过Northern印迹杂交、细胞蛋白代谢标记后免疫沉淀以及Western印迹分析的实验表明，与体内情况类似，在MMH-D3细胞中，视黄醇的存在不影响RBP基因的转录和翻译速率，但对视黄醇结合蛋白的分泌速率调节至关重要。与迄今为止用于类维生素A代谢研究（包括RBP的合成和分泌）的HepG2人肝癌细胞不同，维生素A缺乏在MMH-D3细胞中会导致RBP分泌受抑制以及细胞内蛋白质积累，而视黄醇补充则会迅速导致RBP分泌。该模型在未来关于维生素A在生物体中分布的研究中将非常有用。
