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Macrophages play essential roles in erythrophagocytosis and iron recycling. β-thalassemia is characterized by a genetic defect in hemoglobin synthesis, which increases the rate of iron recycling. We previously showed that reduced expression of the BTB and CNC homolog 1 (BACH1) gene leads to increased phagocytosis of abnormal RBCs by activated monocytes. However, the mechanisms underlying this abnormal RBC clearance remained unclear. Herein, the spleen and bone marrow cells of β-thalassemic mice were examined for erythrophagocytosis CD markers and iron-recycling genes. Higher expression levels of CD47 and CD163 on RBCs and macrophages, respectively, were observed in β-thalassemic mice than in wild-type cells. The decreased expression of BACH1 caused an increase in Nrf2, Spic, Slc40a1, and HMOX1 expression in splenic red pulp macrophages of thalassemic mice. To investigate BACH1 regulation, a macrophage cell line was transfected with BACH1-siRNA. Decreased BACH1 expression caused an increase in CD163 expression; however, the expression levels were lower when the cells were cultured in media supplemented with β-thalassemia/HbE patient plasma. Additionally, the iron recycling-related genes SPIC, SLC40A1, and HMOX1 were significantly upregulated in BACH1-suppressed macrophages. Our findings provide insights into BACH1 regulation, which plays an important role in erythrophagocytosis and iron recycling in thalassemic macrophages.

The regulation of globin gene expression is significantly important to understand the pathogenesis of globin gene disorders. Recent findings have shown that microRNAs (miRNAs, miRs) play an important role in the regulation of globin gene expression. The miR-144 is an erythroid lineage-specific miRNA, in which its expression mediates NRF2 gene silencing and inhibits fetal hemoglobin expression. However, roles of miR-144 to other globin genes expression especially in ɑ-globin cluster remain unknown. This study, thus, examined the functional studies of miR-144 to globin gene expression in K562 human erythroid cell line. The results revealed that ɑ-globin and z-globin gene expression were silenced by the overexpressed miR-144 and that correlated with the reduced expression of KLF1- the suspected target gene. By contrast, transfection with miR-144 inhibitor reversed the silencing effect of miR-144. On the other hand, miR-144 had no effect to β-globin gene expression. Our results sustain the findings of the previous studies that the overexpression of miR-144 correlates with the repressing of NRF2 and 𝛄-globin gene expression. Taken together, our results suggest that miR-144 plays a key role in globin gene expression by silencing 𝛄-globin through NRF2 target mRNA and repressing adult ɑ-globin and embryonic z-globin gene expression possibly by targeting KLF1 gene.