Consequences of DMT1 Mutation on Proliferation and Hemoglobinization of Erythroid Progenitors In Vitro.
Bibliographic record
Abstract
Abstract Disorders of iron metabolism and homeostatis belong to the most common diseases in humans; however, neither the regulation of iron metabolism in erythroid cells nor the pathophysiology of these diseases are completely understood. Last year we reported at this meeting a case of severe hypochromic microcytic anemia caused by a homozygous mutation in the divalent metal transporter 1 (DMT1) gene (DMT1 1285G>C). The defective growth of this patient’s erythroid progenitors in vitro was rescued by the addition of 10 μM iron-salicylaldehyde isonicotinoyl hydrazone (Fe-SIH), an iron chelate that has been shown to deliver iron for heme synthesis without involving the transferrin receptor/DMT1 pathway. To further characterize the impact of the DMT1 mutation on the in vitro growth of erythroid progenitors, 10 μM Fe-SIH, 25 μM hemin or 10 μM zinc chloride (ZnCl2) were added to the erythropoietin (Epo) containing cultures. All substances increased the plating efficiency in normal control as well as mutant BFU-Es. Moreover, all substances led to a 1.4 to 1.6 fold increase in hemoglobin (Hb) content in normal BFU-Es. Although hemin did not significantly increase Hb content in the patient’s BFU-E, the addition of Fe-SIH to the Epo/hemin-containing cultures increased Hb content 2.2 fold. Since Hb synthesis in heme-deficient erythroid cells is inhibited by heme-regulated inhibitor (HRI) kinase, which specifically phosphorylates the α subunit of the eukaryotic initiation factor 2 (eIF2α), we have analyzed the level of eIF2α phosphorylation in cytospine-harvested BFU-Es. Immunofluorescence analysis of erythroid cells with phospo-eIF2α (Ser51) antibody revealed increased phosphorylation of eIF2α in mutant BFU-Es compared to control BFU-Es grown under all aforementioned conditions. The most striking differences were observed in Epo and Epo/hemin containing cultures, in which the fractions of phospho-eIF2α-positive cells were significantly higher in mutant BFU-Es. These data suggest that the block of terminal differentiation/hemoglobinization in the DMT1-mutant erythroid cells is only partially caused by translational arrest of globin chain synthesis; the delivery of iron may be required for other cellular functions apart from Hb synthesis. Interestingly, the most prominent positive effect on the proliferation (i.e., BFU-E cellularity) as well as hemoglobinization of mutant BFU-Es had a combination of Epo/Fe-SIH and ZnCl2; this combination led to the highest increase in Hb content that was associated with almost complete lack of eIF2α phosphorylation. This suggests that the addition of Zn2+ to Fe-SIH containing cultures further stimulates processes that are initially rescued in DMT1-mutant erythroid cells by the addition of iron. We conclude, that the DMT1 defect in erythroid cells is complex, probably involving the combination of transcriptional and translational defects that cannot be rescued by hemin alone. Figure Figure
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".