Mehta Kosha J, Farnaud Sebastien, Patel Vinood B
Department of Biomedical Sciences, University of Westminster, 115 New Cavendish Street, London, W1W 6UW, UK.
School of Life Sciences, Coventry University, 138 James Starley Building, Coventry, CV1 5FB, UK.
Mol Biol Rep. 2017 Oct;44(5):399-403. doi: 10.1007/s11033-017-4123-2. Epub 2017 Aug 24.
In liver hepatocytes, the HFE gene regulates cellular and systemic iron homeostasis by modulating cellular iron-uptake and producing the iron-hormone hepcidin in response to systemic iron elevation. However, the mechanism of iron-sensing in hepatocytes remain enigmatic. Therefore, to study the effect of iron on HFE and hepcidin (HAMP) expressions under distinct extracellular and intracellular iron-loading, we examined the effect of holotransferrin treatment (1, 2, 5 and 8 g/L for 6 h) on intracellular iron levels, and mRNA expressions of HFE and HAMP in wild-type HepG2 and previously characterized iron-loaded recombinant-TfR1 HepG2 cells. Gene expression was analyzed by real-time PCR and intracellular iron was measured by ferrozine assay. Data showed that in the wild-type cells, where intracellular iron content remained unchanged, HFE expression remained unaltered at low holotransferrin treatments but was upregulated upon 5 g/L (p < 0.04) and 8 g/L (p = 0.05) treatments. HAMP expression showed alternating elevations and increased upon 1 g/L (p < 0.05) and 5 g/L (p < 0.05). However, in the recombinant cells that showed higher intracellular iron levels than wild-type cells, HFE and HAMP expressions were elevated only at low 1 g/L treatment (p < 0.03) and were repressed at 2 g/L treatment (p < 0.03). Under holotransferrin-untreated conditions, the iron-loaded recombinant cells showed higher expressions of HFE (p < 0.03) and HAMP (p = 0.05) than wild-type cells. HFE mRNA was independently elevated by extracellular and intracellular iron-excess. Thus, it may be involved in sensing both, extracellular and intracellular iron. Repression of HAMP expression under simultaneous intracellular and extracellular iron-loading resembles non-hereditary iron-excess pathologies.
在肝脏肝细胞中，HFE基因通过调节细胞铁摄取以及在全身铁水平升高时产生铁激素铁调素，来调控细胞和全身的铁稳态。然而，肝细胞中铁感应的机制仍然不明。因此，为了研究在不同的细胞外和细胞内铁负荷情况下铁对HFE和铁调素（HAMP）表达的影响，我们检测了全转铁蛋白处理（1、2、5和8 g/L，处理6小时）对野生型HepG2细胞以及先前已鉴定的铁负荷重组TfR1 HepG2细胞内铁水平、HFE和HAMP mRNA表达的影响。通过实时PCR分析基因表达，用亚铁嗪法测定细胞内铁含量。数据显示，在野生型细胞中，细胞内铁含量保持不变，在低剂量全转铁蛋白处理时HFE表达未改变，但在5 g/L（p < 0.04）和8 g/L（p = 0.05）处理时上调。HAMP表达呈现交替升高，在1 g/L（p < 0.05）和5 g/L（p < 0.05）时增加。然而，在细胞内铁水平高于野生型细胞的重组细胞中，HFE和HAMP表达仅在低剂量1 g/L处理时升高（p < 0.03），在2 g/L处理时受到抑制（p < 0.03）。在未用全转铁蛋白处理的条件下，铁负荷重组细胞的HFE（p < 0.03）和HAMP（p = 0.05）表达高于野生型细胞。HFE mRNA在细胞外和细胞内铁过量时均独立升高。因此，它可能参与细胞外和细胞内铁的感应。在细胞内和细胞外同时铁负荷时HAMP表达受到抑制类似于非遗传性铁过载疾病。
