Expression of Genes Regulating Iron Metabolism in Hepatocyte Cell-Line HepG2 Induced by Sera from MDS Patients.

Abstract

Abstract Liver expression of hepcidin, a key factor in the regulation of iron absorption and recycling, responds commensurately to plasma iron levels. However, in thalassemic humans and mouse models, despite the overt iron overload found, there is a low expression of liver hepcidin and low levels in urine. We have recently shown that hepcidin expression induced by plasma factors in hemochromatosis is overriden in thalassemic patients (Weiser-Stern et al, Brit. J. Haemat.135:129,2006). As low urinary hepcidin levels were also found in patients with Myelodysplastic Syndrome (MDS), mainly in those with refractory anemia (RA) with or without ring sideroblasts (RARS) and iron overload, we set out to examine whether plasma factor(s) in MDS might also affect the expression of liver genes of iron metabolism. Patients and methods. Serum samples were collected from 19 MDS patients, average age: 75.7 y, Hb 6.5–12.0 gm/dl; type and #: RA 11, RARS 5 and 3 RA with excess of blasts (RAEB). The international prognostic score (IPSS) was low in 13/19 and intermediate 1 (Int 1) in 6/19. Only 7/19 patients received less that 5 blood units, while 12/19 received 20–140 units and in 11/19 serum ferritin was >1000 ng/ml. The serum samples were incubated with human hepatoma HepG2 cells, which were harvested and their RNA isolated and analyzed by quantitative RT-PCR for: hepcidin (Hamp), the lipocalin Ngal, HFE, transferrin receptors (TfR) 1 and 2, and DMT1-IRE(−). Values were normalized against those obtained from 3 healthy individuals. Results and discussion. An apparently normal expression of all the above genes was found in 13/19 patients (group I), whereas in the remaning 6/19 (group II) there was: A. a significant increase in expression relative to normal controls (1.0±0.20) of Ngal (4.64±2.15) p<0.01, TfR1 (5.24±0.59) p<0.01, HFE (1.76±0.12) p<0.05 and DMT1-IRE(−) (1.64±0.24) p<0.05; B. a significant decrease in TfR2 (0.19±0.11) p <0.01 and C. an increased Hamp expression in only 3/6 (7.2±5.0) p<0.05 patients and normal expression in 2/6 (1.03±0.3). No clear correlation could be established between expression of the above mentioned genes and iron overload parameters (serum iron and/or ferritin) or the number of transfusions. This study shows a heterogeneous behavior of MDS sera in eliciting gene expression and highlights that increased Hamp expression of HepG2 cells in response to plasma factors is found only in a small fraction of this class of iron overloaded patients. Thus as in thalassemia (1), most of the MDS sera with ferritin levels >1000 ng/ml failed to induce Hamp, suggesting that also in MDS patients there might be circulating factors overriding the serum iron effect on Hamp expression. It remains to be established whether the putative factors modulating expression of key genes of liver iron metabolism in MDS are of the same origin or biochemical character as in thalassemia.