UNIDIRECTIONAL UP-REGULATION OF THE SYNTHESIS OF THE MAJOR IRON PROTEINS, TRANSFERRIN-RECEPTOR AND FERRITIN, IN HEPG2 CELLS BY THE ACUTE-PHASE PROTEIN ALPHA-1-ANTITRYPSIN

Background/Aims: We have previously shown that the hepatic acute-phase protein alpha 1-antitrypsin (alpha 1-AT) is an important mediator of changes in iron metabolism in the course of anaemia of chronic disease , alpha 1-AT profoundly reduces growth of erythroid cells by interferi ng with transferrin-mediated iron uptake, In the present work we inves tigate the effects of alpha 1-AT on hepatic iron metabolism, as the li ver plays a central role in body iron metabolism and in metabolic chan ges during acute-phase response. Methods: The human hepatoma cell line Hep G2 was cultured in RPMI 1640+10% FCS, The effect of alpha 1-AT on transferrin-receptor binding was investigated in equilibrium binding assays with I-125-transferrin. Expression of transferrin receptor was determined by saturation experiments and intracellular ferritin was me asured in cell lysates after incubating cells either alone or with alp ha 1-AT, To determine iron regulatory protein binding activity to iron responsive elements we used gel retardation assays and Northern blot analysis was carried out to investigate transferrin receptor and ferri tin mRNA expression. Results: alpha 1-AT completely prevented transfer rin from binding to its receptor and internalization of the transferri n-transferrin receptor complex on HepG2. In addition, alpha 1-AT cause d a distinct increase in iron regulatory protein binding activity to i ron responsive elements, which is characteristic of iron deprivation, Normally, the synthesis of transferrin receptor and ferritin is regula ted bidirectionally, but alpha 1-AT promoted a unidirectional regulati on, alpha 1-AT increased the synthesis of both transferrin receptor an d ferritin and, moreover, increased cellular amounts of transferrin re ceptor mRNA and ferritin H-chain mRNA. Conclusions: The effect of alph a 1-AT on transferrin receptor synthesis appears to be mediated via ac tivation of iron responsive element binding affinity of iron regulator y protein leading to an increased stability of transferrin receptor mR NA, Changes in ferritin, however, may be related to a transcriptionall y mediated, iron-independent pathway which overrides the influence of activated iron regulatory protein, These specific changes in iron meta bolism are the very ones seen in the course of anaemia of chronic dise ase, Our results emphasize the central role of alpha 1-AT as a mediato r of altered iron metabolism, characteristic of anaemia of chronic dis ease, not only with respect to erythroid cells but also with respect t o liver cells.