Flanagan P R, Hajdu A, Adams P C
Department of Medicine, University Hospital, University of Western Ontario, London, Canada.
Hepatology. 1995 Sep;22(3):828-32.
Iron-responsive element-binding protein (IRE-BP) activity was studied in liver and intestinal samples of hemochromatosis and control patients using a short 32P-IRE-RNA probe on "retardation" nondenaturing polyacrylamide gels. IRE-BP activity was assessed in liver biopsy specimens in 36 patients--16 hemochromatosis homozygotes, 4 hemochromatosis heterozygotes, 6 patients with secondary iron overload, and 10 control patients with normal hepatic iron concentrations. Intestinal IRE-BP activity was assessed in 14 hemochromatosis homozygotes and 16 normal subjects. Endogenous IRE-BP activity was determined from 32P retarded on the gel, and total IRE-BP activity was assessed after reducing tissue samples with 2-mercaptoethanol. Hepatic endogenous IRE-BP activity was inversely related to hepatic iron concentration (r = .59, P < .0002). Mean hepatic endogenous IRE-BP activity in the hemochromatosis homozygotes, 0.25 +/- 0.04 pmol/mg protein, was significantly decreased compared with values in the normal controls, 0.45 +/- 0.06 pmol/mg protein, P < .05. Hepatic total IRE-BP was also significantly decreased in the hemochromatosis patients by gel retardation assay and Western blotting with anti-IRE-BP antibody. Intestinal endogenous IRE-BP activity, total IRE-BP activity, and iron concentration did not significantly differ between hemochromatosis patients and normal control subjects. This suggests that both endogenous IRE-BP activity and the total amount of the protein are downregulated in the liver by tissue iron. Intestinal IRE-BP activity that regulates intestinal transferrin receptor expression is normal in hemochromatosis and appropriate for the intracellular iron concentration.
利用“阻滞”非变性聚丙烯酰胺凝胶上的短32P-IRE-RNA探针，研究了血色素沉着症患者和对照患者肝脏及肠道样本中的铁反应元件结合蛋白（IRE-BP）活性。对36例患者的肝活检标本进行了IRE-BP活性评估，其中16例为血色素沉着症纯合子，4例为血色素沉着症杂合子，6例为继发性铁过载患者，10例为肝铁浓度正常的对照患者。对14例血色素沉着症纯合子和16例正常受试者的肠道IRE-BP活性进行了评估。内源性IRE-BP活性通过凝胶上的32P阻滞来确定，总IRE-BP活性在使用2-巯基乙醇还原组织样本后进行评估。肝脏内源性IRE-BP活性与肝脏铁浓度呈负相关（r = 0.59，P < 0.0002）。血色素沉着症纯合子的平均肝脏内源性IRE-BP活性为0.25±0.04 pmol/mg蛋白质，与正常对照组的0.45±0.06 pmol/mg蛋白质相比显著降低，P < 0.05。通过凝胶阻滞分析和抗IRE-BP抗体的蛋白质印迹法，血色素沉着症患者的肝脏总IRE-BP也显著降低。血色素沉着症患者和正常对照受试者之间的肠道内源性IRE-BP活性、总IRE-BP活性和铁浓度没有显著差异。这表明组织铁在肝脏中下调了内源性IRE-BP活性和该蛋白的总量。调节肠道转铁蛋白受体表达的肠道IRE-BP活性在血色素沉着症中正常，且与细胞内铁浓度相适应。
