Iron deficiency increases DNA-ploidy of megakaryocytic cells and consecutive platelet counts in IBD

Aims: Iron deficiency (ID) is common in IBD affecting up to 90% of patients (Kulnigg S, APT 2006). Additionally, thrombocytosis is a frequent clinical observation that can be reversed by intravenous iron therapy (Gasche C, DDW 2002). Similar to all mammalian cells, the proliferation of megakaryocytic progenitor cells decreases upon ID through cell cycle arrest in G1 (Kulnigg S, DDW 2006), which is inconsistent with the ID-associated increase in platelet counts. Here we show that thrombocytosis also decreases with oral iron replacement, and hypothesize that the increase in platelet counts is rather due to a change in megakaryocyte ploidy (through induction of endomitotic cell cycles) than proliferation.

Methods: Platelet levels in 196 IBD patients who had been treated for ID anemia as part of a controlled clinical trial (Kulnigg S, Am J Gastro 2008) were analyzed before and after 12 weeks of iron replacement therapy with intravenous iron carboxymaltose (FeCarb, n=136) or oral iron sulfate (FeSulf, n=60). The growth and DNA ploidy of megakaryocytic Dami cells were studied under serum-free conditions with (iron rich; IR) or without iron (iron deficient, ID) using Panserin 401 and 401S

(Pan-Biotech, Germany) by MTT assay, and flow cytometry using propidium iodine staining. Iron depletion was assessed by transferrin-receptor expression. Results: In both treatment groups, platelet levels decreased from mean 397 to 317 G/L (FeCarb group, p<0.0001) and from 347 to 309 G/L (FeSulf group, p<0.01); CRP levels remained unchanged. In vitro, Dami cells were cultured 5–7 days in iron-free Panserin to achieve iron depletion, which was confirmed by a 1.4 fold increase in transferrin-receptor expression (range 1.36–1.47). MTT showed lower growth and mitochondrial activity in cells cultured under ID conditions (IR mean fold increase of 7.39±0.15 vs. ID 5.5±0.62, day 6 compared to baseline, p<0.01). In parallel, the fraction of polyploid cells (as defined above 4n) increased (IR 7.7±5% vs. ID 14.5±7.8%, p<0.01). Conclusion: Regardless of the route of iron delivery, platelet levels in this large cohort of IBD patients with ID anemia decreased consistent with the hypothesis of ID enhancing megakaryopoiesis. This seems to be caused by induction of megakaryocytic polyploidy rather than proliferation, indicating a regulatory effect of iron on megakaryocytic endomitosis.